Understanding biological systems is critical to the prosperity, and possibly, survival of the human race. Without it, we are threatened by disease, energy and food insecurity, pollution and climate change.
The COVID-19 pandemic has shown how important it is to have both national and international approaches to research and development with access to the right type of world class equipment, training and expertise.
Structural Biology research included SARS-CoV-2 (COVID-19), snakebite venom, HIV, tuberculosis, malaria, human papilloma virus, cardiovascular disease, as well as equine diseases, and many more. Work has also been done to create industrial enzymes for the manufacture of medicines and commodity chemicals.
“The GCRF START initiative provided an exceptional combination of expertise and experimental resources.”
Cryo-EM structure of Human Papillomavirus (HPV) from data collected at the Electron Bio-Imaging Centre (eBIC) at the UK’s national synchrotron, Diamond Light Source. Image credit and copyright: Melissa Marx and Jeremy Woodward. Created with the UCSF Chimera software package (Pettersen et al., 2004).
Results include 80+ researchers now collaborating from multiple continents, over 50 world class papers published, and a new generation of scientists trained
A virtual event, held on Monday 7 June celebrated the many successes of the Global Challenges Research Fund (GCRF) START grant and the end of the three-year £3.7M grant provided by the UK’s Science and Technology Facilities Council (STFC) from the Global Challenges Research Fund (GCRF) in support of the Synchrotron Techniques for African Research and Technology (START) programme – a unique collaboration between scientists in the UK and Africa, and the UK’s national synchrotron, Diamond Light Source (Diamond). STFC supports the UK’s scientific community by working in partnership with universities, research organisations and government to ensure that researchers have access to large high-quality facilities.
Achievements to date have included more than 50 papers published, with dozens still in the pipeline. Nearly 20 protein structures were deposited in the global Protein Data Bank and over 230 Diamond synchrotron shifts carried out. 30 Postdoctoral Research Assistants/Fellows (PDRA’s/PDRF’s) have been funded, and numerous students introduced to synchrotron science through several workshops, secondments and visits delivered in person and remotely in Africa, the UK and beyond.
The event, hosted by Diamond, chaired by eminent scientist and grant Co-Investigator (Co-I), Professor Sir Richard Catlow, Professor at Cardiff University, University College London and Foreign Secretary at The Royal Society, was attended by START members, funders, and other stakeholders. The event featured case studies, early career impacts, and the importance of Diamond’s collaboration. Research topics included (amongst others), energy materials as catalysts for CO2 hydrogenation to reduce CO2 accelerated climate change; the development and optimisation of thin-film photovoltaic devices (solar cells) for sustainable energy; improving drug design for hypertension and blood pressure (ACE); the world’s first published nitrilaise structure and Africa’s first CryoEM results using Diamond’s eBIC facility. Professor Chris Nicklin, Science Group Leader and Principal Investigator (PI) in the GCRF START grant programme, said,
“START has been an exciting journey catalysed by the GCRF grant which has reaped fantastic results in a remarkably short space of time. By providing the new generation of synchrotron users with access to world class equipment and investing in their skills and capacity, research in the UK and Africa has been enriched and deepened. Going forward, there’s a huge appetite across the START network for a ‘START 2’, especially if the ambition of an African Light Source is to be realised. We are currently looking at ways to continue the momentum and build on START’s promising legacy.”
Dr Thandeka Moyo and Dr Carmien Tolmie are rising stars in the newly emerging Structural Biology network in South Africa. Funded as PDRF’s by the GCRF START grant, their successes stand out in fulfilling the key aims of START, including promoting equality and inclusiveness in science to challenge the under-representation of women. Carmien has made great strides in biocatalysis, investigating enzymes as drug targets for fungal infectious diseases which claim many lives, especially amongst immune-compromised patients. Recently promoted to academic staff at South Africa’s University of the Free State (UFS), she attributes her successes to the mentoring and training she received through the GCRF START grant which funded a secondment to Diamond and the University of Oxford, exposing her to cutting edge scientific techniques such as XChem fragment screening.
Thandeka’s achievements involve notable new biology in HIV (bNAbs) vaccine development projects and Covid-19 research at South Africa’s National Institute for Communicable Diseases. Highly active in mentoring young female scientists and taking part in school outreach activities, Zimbabwean Thandeka encourages the next generation to pursue STEM careers.
Dr Mohamed Fadlalla started out in 2018 as a GCRF START-funded Postdoctoral Research Fellow investigating energy materials at the University of Cape Town (UCT) and has since been promoted into a leadership role as a Research Officer. His story demonstrates how scientific development and scientist career development can work symbiotically, in this case assisted by the GCRF START grant. Mohamed said,
“As well as facilitating amazing science, the GCRF START grant played a substantial role in my career progression as a scientist. This has not only focused my interest into energy materials catalyst development, I have also learnt new energy materials characterisation techniques and conducted experiments at Diamond which were very successful. I started teaching the Catalysis Institute’s MSc. course where I now pass on my new skills to up-and-coming scientists in Africa and have recently been awarded an NRF-Thuthuka grant and UCT block grant to enable my research going forward.”
“If you don’t have this kind of network and support, it is incredibly difficult, for those who are at the start of this field,” said GCRF START Co-I, Professor Michael Claeys, from the University of Cape Town’s Centre for Catalysis and South Africa’s DST-NRF Centre of Excellence in Catalysis, c*change. “It is about building confidence, so this is where the GCRF START grant has played an incredibly important role, improving output and significantly lowering potential hesitancy to use synchrotron techniques, which can seem overwhelming to those new in the field.”
Dr Gwyndaf Evans, START Life Sciences Principal Investigator and Principal beamline scientist on Diamond’s VMXm beamline said,
“It has been rewarding to see the relatively modest investment of time and money can have such a major impact on the sustainability of research expertise, on the development of careers in Africa, on access to large scale facilities around the world, including improving our own systems at Diamond (such as remote access), and to the nurturing of collaborations and networks in South Africa. In Structural Biology, there have been valuable exchanges and collaborations especially XChem laying the foundations for drug discovery work. START is the beginning of embedding the structural research culture in South Africa and other groups around the world. We look forward to what the future holds.”
Funded by the UK’s Official Development Assistance (ODA) Global Challenges Research Fund, the GCRF START grant aims to meet key UN Sustainable Development Goals through Energy Materials for novel devices and improvements in energy efficiency, affordability, and storage (batteries and fuel cells) including renewable energy sources; and Structural Biology to better understand diseases, develop drug targets and vaccines, and drive ‘green’ biotechnology solutions.
Examples of Achievements
More than 230 Diamond synchrotron shifts; over 50 papers published, with dozens in the pipeline; 30 postdoctoral Research Assistants/Fellows (PDRA’s/PDRF’s) funded and numerous students introduced to synchrotron science; 3 ANSDAC workshops and 2 CCP4 workshops sponsored; many crystals and novel structures solved, with more than 20 protein structures deposited in the global protein bank (PDB’s); 8 PDRA’s/PDRF’s trained in Cryo-EM techniques and more than 30 students attended Cryo-EM workshops; PDRA’s involved in the development of new facilities/equipment in the UK and South Africa; 6 fully capable crystallography laboratories equipped across South Africa including in previously under-resourced universities; and multiple workshops, secondments and visits delivered in person and remotely in Africa, the UK and beyond.
Examples of follow on opportunities: Prof. Erick Strauss’ laboratory at the University of Stellenbosch, South Africa, winning a two year AAS Grand Challenges Africa grant focusing on Antimicrobial Resistance and drug discovery; Dr Francis Otieno from the University of the Witwatersrand, South Africa, achieving a British Council Newton Travel grant to conduct experiments on the subject of solar energy with GCRF START’s Co-I, Prof. Moritz Riede, at the University of Oxford; Prof. Dirk Opperman’s group at the University of the Orange Free State winning a Horizon 2020 ERA-NET-Cofund on Food Systems and Climate grant; and various groups able to access to other light sources and neutron sources around the world.
START members hail from the UK, South Africa, Lesotho, Kenya, Egypt, Zimbabwe, Tanzania, Swaziland, Namibia, the DRC, and the Sudan, amongst many other countries.
About the GCRF START grant
The GCRF START grant is a collaborative project that seeks to foster the development of Synchrotron Techniques for African Research and Technology (START). It builds partnerships between world leading scientists in Africa and the UK working together on research using synchrotron science. Funded by the UK’s Official Development Assistance (ODA) Global Challenges Research Fund, the GCRF START grant is delivered by UKRI through Diamond shareholders (the Science and Technology Facilities Council (STFC) and the Wellcome Trust). At the heart of START sits the community of co-investigators whose work in the relevant scientific disciplines is world-leading in their fields. They support a wider group of students and post-doctoral researchers whose contribution to START is vital to nurture future capacity and leadership in the African scientific research community. Working on experiments at the UK’s synchrotron, Diamond, START researchers and students will bring insights to sustainable energy and improvement in health that will have long-lasting legacies across Africa.
For further media information: please contact Diamond Communications: Isabelle Boscaro-Clarke +44 1235 778130
Nicklin, C., Stredwick, R. & Sewell, S. (2022) Synchrotron Techniques for African Research and Technology: A Step-Change in Structural Biology and Energy Materials, Synchrotron Radiation News, 35:1, 14-19, DOI: 10.1080/08940886.2022.2043684
In a bid to design clinical drugs to improve health outcomes for people living with a particular strain of HIV and its mutants, scientists at the University of the Witwatersrand’s Protein Structure-Function Research Unit (PSFRU) in South Africa have embarked on a clinical drug discovery journey with promising results. The aim of the research, which receives funding from the GCRF START grant, is to develop a novel inhibitor specifically designed to target the South African HIV-1 subtype C protease (C-SA PR) and its mutants.
The purpose of developing an inhibitor is to stop C-SA PR’s activity by preventing the formation of mature copies of the human immunodeficiency virus (HIV). This would improve health outcomes in line with Sustainable Development goals across South Africa by increasing drug efficacy, reducing adverse side effects and drug resistance, as well as benefitting populations infected with the HIV subtype C in other sub-Saharan countries, India, China and Brazil[1].
There is no permanent cure for HIV/AIDS which has claimed the lives of an estimated 32.7 million people globally since the beginning of the HIV pandemic (UNAIDS, 2020[2]). A number of drugs have been developed and approved by the USA’s Food and Drug Association (FDA) to increase the quality and duration of life in HIV infected individuals in some parts of the world. However, these are not specific to the HIV-1 protease subtype C which is dominant in South Africa – a country with approximately 20% of the global HIV infection rate and 10.44% of the global AIDS-related deaths (UNAIDS, 2020)[3].
To date, the scientists at the PSFRU have screened ten drugs in the FDA approved and Zinc drug databases, with seven hits that show promise for optimisation as inhibitors. They have solved one protein structure through sophisticated computational modelling, and with high-resolution X-ray crystallography data collected on beamline i03 at the UK’s national synchrotron – Diamond Light Source (Diamond) – have solved the structure of the South African HIV-1 subtype C protease (C-SA PR), the results of which were deposited in the global Protein Data Bank (6I45.pdb) in 2018 (Fig.1).
Fig.1 Ribbon structural representations of the HIV-1 subtype C-SA PR (cyan) and a mutant protease (purple). The structure of the mutant protease was determined using diffraction data obtained at the UK’s national synchrotron, Diamond Light Source (Diamond). PDB ID: 6I45. Sherry, D., Pandian, R., Achilonu, I.A., Dirr, H.W., Sayed, Y. (2018), Crystal structure of I13V/I62V/V77I South African HIV-1 subtype C protease containing a D25A mutation. DOI: 10.2210/pdb6I45/pdb[4]; PyMOL Molecular Graphics System (Schrödinger LLC., Portland, USA[5]).
The research involves characterising the structure and function of the South African HIV-1 subtype C protease (C-SA PR) and its mutants using state-of-the-art computational and experimental methods made possible by the GCRF START grant. The scientists want to understand what role amino acid insertions and mutations the HIV-1 protease may have on clinical drug binding so they can design an effective inhibitor.
This research builds on a previous study[6], in which a blood sample was taken from a drug naive infant born to an HIV positive mother. To prevent the transmission of the virus to her baby, the mother had received reverse transcript inhibitor treatment (but not protease inhibitors) prior to the birth. However, when the baby was born it was found to be HIV positive and a mutation was present rendering current drug therapy ineffective.
“It is the results like this of other research on the South African HIV-1 C-SA PR, and the impact of the disease on individual lives and livelihoods, which drives our motivation. The fact that the baby had developed drug resistance mutations is very rare in mother-to-child transmission but no less concerning,” says Professor Yasien Sayed, who heads up the PSFRU and leads the research on the HIV-1 C-SA PR, “and there is evidence that some adults are also failing drug therapies.We therefore need to develop treatments which work more effectively against the HIV-1 C-SA PR and its mutants if we are going to improve clinical outcomes for the large population of HIV positive adults and children in South Africa, and further afield.”
Some of the first steps in the team’s drug discovery journey have been computational involving Molecular Dynamics Simulation[7] of the HIV wild type C-SA PR and its mutants. After this, drugs in various databases were screened[8]. As a result, the scientists found a promising drug from the FDA database that binds with the HIV wild type C-SA PR and its mutant with best docking scores and energies.
“We modelled the homology structures of the HIV wild type protease and its mutants using the template South African wild type HIV-1 Subtype C Protease (PDB ID: 3U71),” explains Dr Pandian who is a Post-Doctoral Research Fellow funded by the GCRF START grant specialising in the computational aspects of the research. “The structure was solved at 2.72Å using the software packages: Swiss model/modeler/I-Tasser, followed by experimental validation of the modelled structures with in-house computer software. We are excited by the preliminary results, which are better than the current FDA approved drugs, although the computational results now have to be proved through wet lab experiments, along with the best results from the screened from the Zinc database.”
To conduct these studies, the PSFRU has its own computational and wet lab facilities for Molecular Dynamics simulation, docking studies, protein expression and purification. Screening of crystals is carried out using an Oryx8Protein Crystallization Robot (Douglas Instruments, UK) and testing of crystals using a home-source X-ray diffractometer. However, synchrotron facilities are not available on the African continent, so access to the Diamond Light Source synchrotron (Beamlines i03, i04, and also i04-1 and i24) is achieved remotely from the PSFRU lab in South Africa in order to characterise the structure and function of proteins at high resolution.
“Having access through the GCRF START grant to experimental synchrotron techniques like X-Ray crystallography at Diamond to obtain crystals and to solve the structures at high resolutions has been revolutionary for us,” reports Dr Pandian. “It is ultimately the combination of computational and experimental techniques that makes it possible to see how well the drugs are binding to optimise them for the South African HIV-1 subtype C protease (C-SA PR)” [9].
“During the wet lab experiments,” Dr Pandian continues, “we can’t screen the whole drug library for the target protein and it’s very costly to purchase the drugs for screening. The theoretical part of the drug discovery method is therefore useful for generating three dimensional structures for any proteins when the crystal structures are not available in the PDB databases, and for sorting out the best ligand / inhibitors for the protein target before starting protein characterisation wet lab experiments.”
Scientific results are not the only progress being made by the PSFRU team in their research which is meeting the UN’s Sustainable Development Goals for health (SDG 3); great strides have also been made in the PSFRU in terms of education and capacity building in structural biology (SDG 4) with more than 30 postgraduate students involved in the collaboration with the GCRF START grant since 2019. This includes Dr Ramesh Pandian, and Ms Mpho Setshedi who is a MSc. candidate working on the wet lab studies of the HIV-1 C-SA PR and its mutants.
“The research is meaningful,” says Ms Setshedi, “I feel like we are doing a good job and doing something to solve a challenge that impacts South Africa. I hope it contributes something big – an effective HIV inhibitor. In terms of what I am learning, there are challenges in this field but once you get the hang of the techniques you just need to persevere. Getting funding is a struggle in my field generally and there aren’t a lot of women doing this work. There have also been challenges caused by the COVID-19 lockdown in 2020 – but I haven’t let anything discourage me.”
[7]The Molecular Dynamics (MD) simulations of validated structures were performed under physiological conditions for 100ns using GROMACS software packages. The MD simulated structures were analysed thoroughly and extracted the energy minimised structure for further analysis. Important parameters such as root mean square deviation (RMSD), root mean square fluctuation (RMSF), radiation of gyration (Rg) and hydrogen bonding analysis were carried out.
[8]The energy minimised structures of HIV wild type and its mutants were used for the screening of drugs from different data bases such as the Zinc database and FDA approved drugs database.
[9]The results of the binding pocket analysis of the protease complex form obtained by the docking studies with best ligand directed the scientists to modify the side chain with the combination of different R group of the drug to improve the binding affinity.
On International Women’s Day (today – 8th March), we highlight the achievements of Melissa Marx, an MSc medical biochemistry student in the University of Cape Town’s (UCT) Faculty of Health Sciences (FHS), who is conducting ground-breaking research in the field of cervical human papillomavirus (HPV) with the assistance of the GCRF START grant. HPV is the cause of most cervical cancer cases amongst women worldwide. Melissa’s research into HPV, which forms part of a larger departmental programme, was funded by the GCRF START grantwhich provided access to the UK’s world class national synchrotron, Diamond Light Source.
Marx’s research focused on visualising the effect of an enzyme found within the reproductive tract on the structure of the virus, as it occurs during the infection process. To achieve this, she and fellow researchers studied HPV pseudoviruses (non-infectious and synthetic viruses) using laboratory-based techniques, structural biology and computational work.
Melissa hopes that her findings will set a strong foundation as scientists work towards discovering preventative and therapeutic options for HPV infection to decrease the high burden of HPV infection in South Africa, on the continent and around globe. Cervical cancer is the fourth most common cancer among women worldwide.
“Without the help of the GCRF START grant, much of my research would not have been possible. The grant enabled me to apply cutting-edge structural biology techniques to gain insights into the structure of HPV,” she said.
“I was also incredibly fortunate to have collected data at the Electron Bio-Imaging Centre at Diamond Light Source in the United Kingdom. This was only possible with funding from the GCRF START grant.”
Melissa Marx, MSc medical biochemistry student in the University of Cape Town’s (UCT) Faculty of Health Sciences. Melissa’s research in the field of cervical human papillomavirus (HPV) is assisted with funding by the GCRF START grant. Photo supplied by Melissa Marx.
And why collaborating with young female scientists in Africa is reaping great results.
Timed to coincide with UN International Day of Women and Girls in Science on 11 Feb, and to inspire more female students to study and work in science, the GCRF START grant has announced the results of its three year project launched in March 2019. To date it has directly collaborated with nearly 50 young African research students and given access to almost 100 synchrotron beamline sessions. Over half of START’s students are female scientists who are demonstrably changing perceptions and increasing the possibilities for women choosing long term STEM research careers.
“Globally UNESCO figures show that only 30% of researchers are female and they occupy only 20% of STEM leadership positions. These figures are even lower in many countries in Africa underlining how important it is to challenge women’s under-representation. Young female African scientists are vital both for their research and as role models and mentors for the next generation. So we are really delighted to see many of the young women we collaborate with through the START grant, making great strides and achieving some incredible results in the fields of structural biology and energy materials”
Prof. Chris Nicklin, Science Group Leader and Principal Investigator (PI) in the GCRF START (Synchrotron Techniques for African Research and Technology) grant programme.
GCRF START is an innovative collaboration between Diamond Light Source, the UK’s national synchrotron, and higher education and research partners in the UK and Africa. It is funded by the Science and Technology Facilities Council under the UK government Global Challenge Research Fund programme. It is enabling and inspiring researchers from this, and the next generation of Africans to choose careers in science and find African and joint UK-African solutions to some of the world’s most pressing health and environmental challenges. A key goal is to challenge the under-representation of women in science by providing access to world-class scientific facilities, funding, training, mentoring, and unique international collaborations. Great results have been achieved in a relatively short space of time because START scientists get access to specialist technologies and facilities not available on the African continent – like beamtime on the Diamond synchrotron.
One indicator of the success of the programme is how the tiny community of structural biologists in Africa has grown across South Africa including a whole new generation of women. Similarly, in energy materials, the gender factor has traditionally been a barrier, so having young women entering materials science is great progress. Additionally, all these women participate in outreach and act as role models to inspire girls to choose STEM careers. Female START successes include:
Priscilla Masamba has solved the partial structure of a protein from Schistosoma mansoni, a parasite responsible for the debilitating disease Schistosomiasis (Bilharzia) which is endemic in more than 78 countries, with an estimated 4 million people infected in South Africa alone. Her work will contribute to drug discovery efforts and is notable because she was the first student from the University of Zululand, South Africa, to use the Diamond synchrotron, which she did remotely from a lab in South Africa learning many scientific techniques for the first time;
Thandeka Moyo is part of a leading South African team working on HIV/AIDS vaccine research and is currently researching Covid-19; Originally from Zimbabwe, Thandeka mentors early career female scientists and is a role model for school children;
Gugulethu Nkalais investigating new generation renewable energy storage systems in South Africa to help close the energy poverty gap; she is active in inspiring girls into STEM;
Lizelle Lubbe is a GCRF START grant-funded Postdoctoral Research Fellow in Structural Biology (a scarce skill in Africa). Lizelle is one of only a handful of scientists in Africa as a whole trained in single particle cryo-EM – a cutting-edge technique for determining the structure of proteins;
Michelle Nyoni is studying energy materials to improve the performance of Lithium-ion batteries for portable electronics and renewable energy sources to make them affordable and improve their environmental footprint to tackle climate change. Michelle is also a chemistry lecturer in Zimbabwe.
“The GCRF START grant has been a game-changer for young African scientists, particularly from under-represented groups such as female, and black scientists, enabling them to enter the fields of Structural Biology and Energy Materials and thrive.”
GCRF START Co-Investigator, Prof. Edward D. Sturrock from the University of Cape Town, South Africa.
One young scientist working with START, Gugulethu Nkala, is an Energy Materials PhD student from South Africa. The eldest of three daughters and first in her family to go to university, she remarks; “Seeing a black girl in science, makes girls see that there is someone, just like them, who has gone this far. We are breaking barriers that makes science seem unattainable, by being the link between science and society, made possible by funding bodies like the GCRF START grant.”
Access to inclusive quality education and lifelong learning opportunities is a distant dream for many young people across Africa, especially women. Few have the opportunity to finish school, let alone reach university to study world-class science, be mentored by experts or continue to postdoctoral studies. This can be due to lack of access to resources at home institutions, insufficient grant writing experience, lack of mentors or supervisors, inadequacy of facilities, and poor postdoctoral pay.
“It is important to support and mentor young women in science especially since women are largely under-represented, particularly in the case of the physical sciences, the field in which we work. I found that having access to synchrotrons and also building international collaborations through the GCRF START grant programme has not only allowed the young women that I work with to gain better skills but they also grow in confidence about their abilities”
Professor Caren Billing, Energy Materials Research Group Co-Principal Investigator (Co-PI), Lecturer and Associate Professor in the School of Chemistry at the University of the Witwatersrand, South Africa.
The GCRF START grant supports young scientists working on key Climate, Energy, Health, and Education challenges in line with the UN’s Sustainable Development Goalsby building partnerships between world leading scientists in Africa and the UK and enabling them to work together on research using synchrotron science. The project focuses on developing and characterising new energy materials, for example in the development of solar cells or improving energy efficiency through novel catalysts, and structural biology to understand diseases and develop drug targets for better treatments and potential vaccines. The START programme is grant-funded through the UK’s Global Challenges Research Fund (GCRF) and delivered by UK Research and Innovation (UKRI) through the Science and Technology Facilities Council (STFC) and the UK’s national synchrotron facility, Diamond Light Source.
“Science is a collaborative discipline. Yet science is being held back by a gender gap. Girls and boys perform equally well in science and mathematics – but only a fraction of female students in higher education choose to study sciences. To rise to the challenges of the 21st century, we need to harness our full potential. That requires dismantling gender stereotypes. It means supporting the careers of women scientists and researchers.” United Nations, Secretary General, Antonio Guterras, United Nations International Day of Women and Girls in Science – 11 February
We are delighted to announce that our exciting Sci-Art collaboration with The Keiskamma Trust in South Africa is well underway! In the spirit of Ubuntu [1], funded by the GCRF START grant, the collaboration involves a unique series of tapestries inspired and created by community-based artists and crafters from the Trust’s flagship Keiskamma Art Project located in South Africa’s Eastern Cape. The artwork is based on concepts provided by scientists in the UK and Africa working on GCRF START-related Energy Materials and Structural Biology themes. The aim of the collaboration is to stimulate shared learning and dialogue on solutions to local and global challenges in line with Sustainable Development Goals: from alternative energy solutions to tackle pollution and climate change, and biotechnology for food security, to improved health outcomes through novel drug discovery and design.
Embroidery crafters from The Keiskamma Trust’s flagship Art Project sewing tapestries depicting GCRF START-related research by scientists in the UK and Africa. Photo credit: Pippa Hetherington, The Keiskamma Trust. Copyright: The Keiskamma Trust
The Keiskamma Trust is a small, Non-Profit Organisation (NPO) in South Africa’s Eastern Cape dedicated to addressing HIV/AIDS and poverty holistically through health, art, music and education initiatives. With unemployment levels in the region’s rural areas up to 90%, and water shortages, poor nutrition, lack of electricity, and diseases like HIV/AIDS, Tuberculosis (TB) and diabetes highly prevalent, hardship is an everyday experience for the remote communities the Trust serves. The artists and crafters creating the tapestries hail from the villages of Hamburg, Bodiam and Bell in the surrounding communities. Overcoming many challenges including months of Covid19 lockdown, they have already completed several panels, including one large panel and a series of smaller ones, with more panels nearing completion.
While the large ‘Our Vision for Africa’ panel gives vibrant expression to a community-led vision of a future with clean air and access to sustainable energy and good health, the smaller panels intricately depict a host of specific research topics. These range from explaining the purpose of the UK’s national Diamond Light Source synchrotron (Diamond), which lies at the heart of the GCRF START grant, to research on structures of proteins for novel HIV, blood-pressure and anti-fungal infection drug treatments, studies exploring organic solar energy materials, and themes on the role of catalysis, which underpins food production, generation of clean energy, and maintenance of clean water and air.
The ‘Our Vision for Africa’ large panel designed by Siyabonga Maswana and Sanela Maxengana, artists of the Keiskamma Art Project. This panel depicts the artists’ vision for the future of their village, which is what the GCRF START grant is all about: access to sustainable development goals of good health, sustainable energy, and a clean environment. Each element on the panel relates to different aspects of research from catalysis and energy materials, to drug discovery and biotechnology. Photo credit: Jeremy Woodward. Copyright: University of Cape Town on behalf of a collaborative project with the Synchrotron Techniques for African Research and Technology (START) in the United Kingdom funded by the Global Challenges Research Fund (GCRF) START grant. ‘Our Vision for Africa’, 2020, Keiskamma Art Project. Embroidered by Nomgcobo Nompunga and Asanda Nompunga.
Artwork commissions like the SciArt collaboration bring much needed employment and income generation opportunities to the staff employed by the Keiskamma Art Project. Such commissions can also provide a platform for empowering skills development and learning, as Cebo Mvubu, theproduction managerat the Keiskamma Art Project, explains, “Here in South Africa the unemployment is too high and now even more with the Covid19 crisis”, says Cebo. “Commissions like the GCRF START Sci-Art project help many people from our villages, directly and indirectly. The income feeds our families and helps send our kids to school, and even if just one person is working on a project like the START commission, this helps support more than 5-8 people in their extended family. We also benefit from the skills we learn when doing these commissions and the publicity the project gets.”
The villages served by The Keiskamma Trust are located in a remote, rural part of South Africa’s Eastern Cape. Photo credit: Pippa Hetherington, copyright: The Keiskamma Trust.
There are many stories of hope among the crafters and artists at the Keiskamma Art Project, not least those reflecting the strength of the women in the region – described as ‘elabafazi’ in the local Xhosa language. One such story is from a crafter who is the sole breadwinner in her family, despite suffering from chronic health conditions, she alone supports her younger sisters, brothers and her daughter who are all unemployed.
“I have a big challenge because I am a single mother and I have to look after the kids at home,” the crafter from the Keiskamma Art Project explains. “I am not 100% health-wise so it is a big thing to feed everyone, enable them to go to school, and to keep a home. But since the year 2000, when I was accepted by the The Keiskamma Trust, I have employment which supports my family. I have learnt sewing, drawing, embroidery, felt-making, screen printing, pottery, and painting. It is a big opportunity to be the part of START’s Sci-Art project.”
A crafter from the Keiskamma Art Project sewing a tapestry panel for the GCRF START grant’s Sci-Art project depicting research on blood pressure. Photo credit: Cebo Mvubu, The Keiskamma Trust. Copyright: The Keiskamma Trust
There is a strong desire amongst the artists and embroidery crafters working on START’s SciArt commission for dialogue with the scientists in order to learn more about the concepts behind their creations, especially what the science could mean for their families and communities, “We want to learn from each other and from the scientists as we see this as a collaboration,” says Cebo. “It is one of the things we would love to know: what the scientists do. For example, we would like to know more about solar energy. We need new energy options here because we have little electricity and we often have load-shedding. But we do have lots of sunshine! We also hear that the scientists want to learn from us too. We would like to show them how we do this artwork.”
Cebo Mvubu, Production Manager at The Keiskamma Trust’s flagship Keiskamma Art Project. Photo credit: The Keiskamma Trust. Copyright: Diamond Light Source
“There is a hunger here for greater connectivity. We can fashion a bridge from science to art through the interpretation of research concepts into embroidery, but it would mean so much to us if the science itself could touch our communities,” says Michaela Howse, curator and manager at the Trust’s Keiskamma Art Project. “It is rare that science being done in metropolitan centres, or internationally, reaches our villages and it is very exciting to work on concepts that seem to come from another world,” explains Michaela. “Dialogue between artists in our unique contexts, and the GCRF START scientists in theirs, may enrich both parties. Perhaps the applications of the research might one day grow to directly impact the needs of communities whose main concerns remain improved health, education opportunities, as well as dignity in life above all things.”
To encourage dialogue, the artists and embroidery crafters have recently written a letter to the scientists collaborating with START asking them to share how the concepts provided by the researchers could impact their villages. In the letter they ask: “Can your work educate us or help us in understanding energy, electricity, water, disease, better health and better lives? We would love to hear from you.”
The scientists are currently responding with letters of their own, explaining what the science might mean for improving everyday lives. In one such letter, scientists collaborating with START from the Biocatalysis and Structural Biology Research Group at the University of the Free State, in South Africa, respond by citing the inspiration behind their research to find new anti-fungal compounds which can be used in the health and agricultural sectors, “Dear Keiskamma community,” the letter states. “Thank you for your letter – both your commitment to your art and your determination in difficult circumstances inspire us to work hard on our research and make a difference in people’s lives.”
“I find it motivating to see the artists represent their vision of a better future through their artwork and I hope the science that I do has a positive impact on the world,” says GCRF START Co-Investigator, Dr Jeremy Woodward, who is the Principal Investigator in the Structural Biology Research Unit at the University of Cape Town. “These artworks show what the GRCF START grant is all about: using the most sophisticated technology in the world to enable Africans to solve African and global problems. I particularly hope that young people see these artworks and this plants the seed so that people see that science can be done anywhere in the world, by anyone.”
“It has been a great joy to see our science come to life through the eyes of the Keiskamma artists and I am very excited about the project’s potential to uplift communities in Africa,” says GCRF START Postdoctoral Research Assistant, Dr Lizelle Lubbe, from the University of Cape Town. “I hope this collaboration will break down barriers to science and inspire future generations of researchers and innovators, as well as stimulate dialogue with the communities impacted by the challenges that people in Africa and around the world face.”
The ‘Angiotensin Converting Enzyme’ panel. A tapestry designed by Cebo Mvubu of the Keiskamma Art Project based on research by Prof. Ed Sturrock (GCRF START Co-I) and GCRF START Postdoctoral Research Assistant, Dr Lizelle Lubbe, from the University of Cape Town, South Africa. The panel shows a snake wriggling through a blood vessel that has become affected by a build-up of fats, cholesterol and calcium (atherosclerosis). High blood pressure is a major cause of atherosclerosis and can lead to heart attacks and strokes. Certain snake venoms contain compounds that, when injected, cause their prey to lose consciousness from a drop in blood pressure. The venom of the Brazilian viper inhibits angiotensin converting enzymes and forms the basis for medicines that are used to lower blood pressure and treat heart disease.
Photo credit: Jeremy Woodward. Copyright: University of Cape Town on behalf of a collaborative project with the Synchrotron Techniques for African Research and Technology (START) in the United Kingdom, funded by the Global Challenges Research Fund (GCRF) START grant. ‘Angiotensin Converting Enzyme’, 2020, Keiskamma Art Project.
Embroidered by Nosiphiwo Mangwane.
The ‘Flexible Solar Cells’ panel. A Solar Energy tapestry designed by artist Nozeti Makubalo from the Keiskamma Art Project. The panel design is based on a collaborative concept provided by Prof. Moritz Riede (GCRF START Co-I) and Postdoctoral Research Assistant, Dr Pascal Kaienburg, from the University of Oxford, and Prof. Chris Nicklin (GCRF START PI) and Postdoctoral Research Associate, Dr Thomas Derrien, from the UK’s national synchrotron, Diamond Light Source (Diamond), all of whom work closely together on Solar Energy research. The materials being studied can be used to make solar cells which harness the sun as a source of energy. The research looks at how to improve the efficiency of materials in ‘organic semiconductors’ to make them commercially viable. These are more lightweight, flexible, environmentally friendly, and easier to deploy in rural environments than heavy, stiff panels of silicon-based solar cells. The data obtained tells us how these materials organise themselves on devices, which can affect how well the solar cells work.
Photo credit: Jeremy Woodward. Copyright: University of Cape Town on behalf of a collaborative project with Synchrotron Techniques for African Research and Technology (START) in the United Kingdom funded by the Global Challenges Research Fund (GCRF) START grant. ‘Flexible Solar Cells’, 2020, Keiskamma Art Project.
Embroidered by Nozeti Makubalo.
The ‘Aspergillosis’ panel. A tapestry designed by artist Siyabonga Maswana from the Keiskamma Art Project based on a concept provided by Dr Diederik Opperman (GCRF START Co-I) from the University of the Free State’s Biocatalysis and Structural Biology Research Group in South Africa. Opportunistic fungal pathogens (agents) invade vulnerable individuals, such as immune-compromised patients, and cause life-threatening health conditions (mucoses). Anti-fungal agents are used to combat mycoses but current therapies often suffer from toxicity, as well as emerging anti-fungal resistance, prompting the search for alternative medicinal drug targets. The panel depicts invasive aspergillosis, an infection caused by a type of fungus, growing on the lungs. The bright light of the UK’s synchrotron, Diamond Light Source, and a technique called X-ray crystallography are used to examine the structures of fungal redox enzymes (special types of proteins) as novel anti-fungal drug targets.
Photo credit: Jeremy Woodward. Copyright: University of Cape Town on behalf of a collaborative project with Synchrotron Techniques for African Research and Technology (START) in the United Kingdom funded by the Global Challenges Research Fund (GCRF) START grant. ‘Aspergillosis’, 2020, Keiskamma Art Project.
Embroidered by Nomakhaya Dada.
‘Chemistry from Plants’ panel. A tapestry designed by artist Siyabonga Maswana from the Keiskamma Art Project based on a concept provided by Dr Jeremy Woodward (GCRF START Co-I) from the University of Cape Town’s Structural Biology Research Unit in South Africa. Plants produce a variety of chemical compounds to defend themselves from being eaten and these poisons need to be detoxified by the plant when not needed. The panel depicts a small weed – Red Shepherd’s Purse – which repels insects by producing poisonous compounds called nitriles. These are broken down by three different enzymes, each converting nitriles of a different size. How these enzymes worked was a mystery until now because we couldn’t visualise them. Normally, enzymes arrange themselves into crystals that allow us to determine the positions of every atom but in this case it wasn’t possible because of their pentameric shape, as shown on the panel by pentagons that do not assemble into a space-filling pattern. Now, using the UK’s Diamond Light Source Synchrotron (Diamond) and the Titan Krios III (beamline M06) at Diamond’s Electron Bio-Imaging Centre (eBIC), Dr Woodward has been able to image these enzymes for the first time, paving the way to design new enzymes for a range of ‘eco-friendly’ biotechnology applications, from cleaning up toxins in contaminated land to improving crop types and yields, and helping design medicines with fewer side effects.
Photo credit: Jeremy Woodward. Copyright: University of Cape Town on behalf of a collaborative project with Synchrotron Techniques for African Research and Technology (START) in the United Kingdom funded by the Global Challenges Research Fund (GCRF) START grant. ‘Chemistry from plants’, 2020, Keiskamma Art Project.
Embroidered by Thembisa Gusha.
Typical village scene in the rural communities of the Eastern Cape of South Africa. Photo credit: Cebo Mvubu. Copyright: The Keiskamma Trust
Tapestry topics
Catalytic CO2 conversion to methanol for producing renewable and sustainable fuels; new compounds for controlling blood-pressure; enzymology for solutions to food security; anti-fungal drug targets for life-threatening fungal infections (mycoses); the structure of the South African HIV-1 Subtype C Protease for insights into a possible HIV vaccine/treatments; research into antibiotic-resistant strains of the bacteria Staphylococcus aureus; improving efficiency of solar cell materials; finding solutions to diseases like Malaria, and research on Rotaviruses which are the most common cause of diarrheal disease.
South Africa: University of the Witwatersrand; University of Cape Town; Stellenbosch University; University of the Free State; North-West University; Aim Shams University; University of Limpopo; University of Pretoria; National University of Lesotho; National Institute of Communicable Diseases (NICD).
United Kingdom: Diamond Light Source; University of Oxford; University of Southampton; University of Cardiff; University of Sheffield.
More about The Keiskamma Trust
The Keiskamma Trust uses art and heritage/tourism to alleviate long-standing poverty and unemployment in the communities of Hamburg, Bodiam and Bell in the Eastern Cape of South Africa. Founded in 2000 by the local Xhosa community with the help of the Trust’s first director, artist and doctor – Dr Carol Hofmeyr – the Trust’s community driven and inspired Keiskamma Art Project works to develop creative skills to empower mainly women and young members of the community. It does this through turning inherent talents into sustainable income-generating activities, showcasing the local culture and heritage, and aiding the archiving of the Eastern Cape rural collective memory and preservation of oral history. Read more here about The Keiskamma Trust. Related articles: The Keiskamma Art Project: Restoring Hope and Livelihoods:https://www.tandfonline.com/doi/abs/10.1080/00043389.2017.1338648?journalCode=rdat20
Contacts
For more information about the GCRF START SciArt project, please contact : Dr Jeremy Woodward
[1] ‘Ubuntu’ or ‘umntu ngumntu ngabantu’ in the isiXhosa language means ‘I am because you are’. In the Oxford Dictionary and Oxford Learners’ dictionary respectively, Ubuntu is defined as a quality that includes the essential human virtues of compassion and humanity or ‘the idea that people are not only individuals but live in a community and must share things and care for each other.’
“We have been privileged to have worked with community members who are so committed to the research that could one day realize our shared vision of a world without AIDS.”
Since HIV was found to be the cause of acquired immune deficiency syndrome (AIDS) in 1983, scientists have been working endlessly towards the development of an effective vaccine to end the global HIV pandemic which has claimed more than 32 million lives[1] and impacted millions more. Unfortunately, the best vaccine candidate we have had to date was from the famous Thai RV144 trial which only resulted in 31.2% efficacy (Rerks-Ngarm et al., 2009[2]). However, hope remains and more than 30 years after the discovery of HIV, we have uncovered many vulnerabilities of the virus which could lead to the development of an effective HIV vaccine to solve one of the big global challenges of our age.
One of the keys to a successful vaccine is the use of broadly neutralizing antibodies (bNAbs). These special antibodies bind to the HIV envelope protein and prevent the virus from infecting host cells. What makes them even more special is that they can bind numerous mutated versions of the virus and therefore overcome the problem of the HIV variability. Understanding the diverse ways that antibodies use to target the HIV envelope is important to the development of an HIV vaccine which can produce such unique and unusual antibodies and effectively protect vaccinated individuals from HIV infection.
My name is Dr Thandeka Moyo and I am a GCRF START grant-funded Postdoctoral Research Fellow at South Africa’s National Institute for Communicable Diseases (NICD), affiliated to the University of the Witwatersrand (Wits). Over the past few years, my colleagues and I have gained new insights into bNAbs in chronic HIV infection, insights which contribute significantly to the worldwide hunt for an HIV vaccine. Most recently, with access to the UK’s national synchrotron – Diamond Light Source (Diamond) – facilitated by the GCRF START grant, we were able to solve the structure for one member of a family of antibodies which has revealed a uniquely long loop in the light chain of the antibody – a loop up to three times longer than other published anti-HIV antibodies![3] Such insights are exciting, providing opportunities not only to expand my skills and knowledge as an early career scientist but also to inspire further hope that we will one day have all the necessary information to design an effective vaccine to end the global HIV pandemic.
South Africa has the biggest HIV epidemic in the world, with an estimated 7.5 million people infected nationally according to UNAIDS[4]. In this article, I will outline some of the insights achieved over several years of investigating broadly neutralizing antibodies through women participating in the CAPRISA Cohort – a research programme based in the KwaZulu-Natal province of South Africa. Without these women enabling us to study their donated samples, we would still have many unanswered questions.
“The experiences of the CAPRISA bnAb cohort studies epitomize the strength of the relationship between CAPRISA researchers and the local communities – a relationship of respect and equality.”
The rural town of Vulindlela in KwaZulu-Natal, South Africa, where the CAPRISA Vulindlela Clinical Research Clinic is based. Photo credit: Dean Demos
Investigating broadly neutralizing antibodies in chronic HIV infection
A subset of individuals who are infected with HIV develop broadly neutralizing antibodies (bNAbs) in chronic HIV infection. Unfortunately, these special antibodies are very unusual with characteristics not found in most other antibodies that we have in our bodies. For example, these antibodies are highly mutated and have longer “arms” that reach out and bind to the virus. Most antibodies do not have these characteristics, so researchers have spent years studying these unique bNAbs to get a better sense of how to produce them with an HIV vaccine.
In our laboratory at the NICD, we study antibodies in HIV-infected women from KwaZulu-Natal who participate in the CAPRISA Cohort. Established in 2003, this Cohort has tracked women over several years from before HIV infection, through to when a subset of them were just infected (acute stage), and years after infection (chronic stage). Throughout the course of the study, the women received healthcare and HIV counselling through the Cohort and have continued to participate in it for years. Blood samples were taken at various time points and from these samples we have been able to track the evolution of the viruses in these women as well as the way their antibodies have adapted throughout infection – with some women developing these special bNAbs.
The CAPRISA Cohort has been invaluable in providing us with novel information on how bNAbs develop as the virus mutates, as well as how we can engineer a vaccine strategy more widely to make these antibodies. This has helped us understand the development of bNAbs and how we can use these antibodies for an effective HIV vaccine. Outlined below are examples from three women in the Cohort (referred to as CAP256, CAP248 and CAP314 respectively) who have developed special antibodies.
CAPRISA Cohort participants CAP256 and CAP248
CAPRISA Cohort participant CAP256 became infected with HIV during the study and then re-infected with another variant form of HIV 15 weeks after initial infection – a phenomenon referred to as superinfection. A related virus to the superinfecting virus changed the immune response in this woman and she developed bNAbs after this event. Scientists in the USA isolated antibodies from the blood donated by CAP256[5] and discovered that the best antibody isolated bound to the apex (the top region) of the HIV envelope protein. This antibody is the most potent antibody isolated to date that binds to this target and the exact mode of binding for this antibody was not understand until recently. Led by our collaborators at the National Institute of Health in the USA, a study of the high-resolution structure of this antibody bound to the HIV envelope using cryo-electron microscopy revealed that it uses two distinct mechanisms to bind to this region (Gorman et al., 2020[6]). The use of these diverse strategies is likely the cause of its extremely high potency. The antibodies from CAP256 are currently in an HIV prevention clinical trial with results expected in the next few years.
Another CAPRISA Cohort participant, CAP248, also developed bNAbs. Researchers isolated an antibody from this participant which was unusual in the HIV envelope site that it targeted. Using negative stain electron microscopy (Scarff et al., 2018[7]), they showed that this antibody from CAP248 bound to a target proximal to the viral membrane and parts of the antibody interacted directly with viral membrane (Wibmer et al., 2017[8]). This mode of binding is unique and represents a novel way an antibody can bind to the HIV envelope protein. This novel binding mechanism may provide insight into the design of an HIV vaccine candidate that can produce this type of antibody response.
Solving a unique antibody structure with the help of the GCRF START grant – CAPRISA Cohort participant CAP314
The last participant to highlight in this article is CAP314. CAP314 developed bNAbs within two years of infection which is a relatively short time for HIV-infected individuals to develop these special antibodies. We isolated antibodies from three families of antibodies that developed in this individual. One family of antibodies mutated over time in response to the mutating HIV variants circulating in CAP314 at the same time. We were able to solve the structure for one member of this antibody family by X-ray crystallography on Diamond’s i03 beamline, in 2019, with Dr Dave Hall as our local contact in the UK for the beamline session and access provided by the GCRF START grant. Solving the structure of this antibody has revealed its uniqueness in that it has an extremely long light chain loop. The loops of antibodies are like arms which reach out and attach to their target on the HIV Envelope. Most anti-HIV antibodies have long loops on the heavy chain of the antibody, but this antibody has a uniquely long loop – up to three times longer than other published anti-HIV antibodies – in the light chain of the antibody. This long light chain loop reaches into the HIV envelope protein and makes the necessary contacts for this antibody to bind to the virus and stop it from infecting cells. Novel and unique binding mechanisms of antibodies like these give us important insights which could help us in our quest to design an HIV vaccine.
Global solidarity, shared responsibility through phenomenal CAPRISA Cohort and collaborations
We are very grateful to the women in the CAPRISA Cohort who make our vital work towards the goal of HIV prevention possible, and to our collaboration with the GCRF-funded START grant. START is a phenomenal initiative supporting capacity development of structural biologists throughout Africa by providing improved access to world class synchrotron equipment, mentoring and expertise. I have been fortunate to have found extremely supportive mentors in START Co-Investigator’s (CoI’s), Prof. Penny Moore and Prof. Lynn Morris, who have encouraged my independence and supported me throughout my Postdoctoral studies.
“Over the last decade, we have learned an incredible amount about how some HIV infected women make broadly neutralizing antibodies. These insights have significantly contributed to HIV vaccine design. This has only been possible because of the extraordinary commitment shown by CAPRISA Cohort donors who come back again and again, and the clinical staff who care for them. We are truly indebted to them.”
GCRF START Co-I, Prof. Penny Moore, University of the Witwatersrand and the National Institute for Communicable Diseases.
The UN states that a core principle of the 17 Sustainable Development Goals (SDGs), and of the AIDS response, is that “no one should be left behind.“ The AIDS epidemic cannot be ended without “the needs of people living with and affected by HIV, and the determinants of health and vulnerability, being addressed.”– UNAIDS[9]
Read more here about HIV/AIDS and the UN Sustainable Development Goals.
World AIDS Day 2020 – Global solidarity, shared responsibility. Photo credit: UNAIDS
More about Dr Thandeka Moyo
GCRF START Postdoctoral Research Fellow, Dr Thandeka Moyo, holds a BSc with distinctions in Biochemistry and Microbiology and a BSc (Hons) in Biochemistry from Rhodes University. She went on to obtain a MSc (Med) and PhD in Clinical Science and Immunology from the University of Cape Town where she looked at the mechanisms used by various HIV strains to gain resistance to broadly neutralizing antibody responses. Based at South Africa’s National Institute for Communicable Diseases and affiliated to the University of the Witwatersrand, Thandeka’s postdoctoral research involves understanding the structure and function of HIV neutralizing antibodies by X-ray crystallography. More recently she has added SARS-CoV-2 to her research focus, developing serological assays to measure humoral responses to infection and vaccination.
GCRF START Co-I, Prof. Penny Moore, is a Reader and DST/NRF South African Research Chair of Virus-Host Dynamics at the University of the Witwatersrand and the National Institute for Communicable Diseases. She holds a joint appointment as Honorary Senior Scientist in Virus-Host Dynamics at the Centre for the AIDS Programme of Research in South Africa (CAPRISA), University of KwaZulu-Natal. Moore co-directs a team of more than 15 scientists and 10 graduate students, with the team’s research focused predominantly on HIV neutralizing antibodies and their interplay with the evolving virus.
[2] RERKS-NGARM, S., PITISUTTITHUM, P., NITAYAPHAN, S., KAEWKUNGWAL, J., CHIU, J., PARIS, R., PREMSRI, N., NAMWAT, C., DE SOUZA, M. & ADAMS, E. 2009. Vaccination with ALVAC and AIDSVAX to prevent HIV-1 infection in Thailand. New England Journal of Medicine, 361, 2209-2220; doi: 10.1056/NEJMoa0908492.
[3] The structure was solved by Dr Thandeka Moyo from the NICD, South Africa, and Taylor Sicard and Dr Jean-Philippe Julien from the University of Toronto, Canada.
[5] The donor was identified by Prof. Penny Moore in research that commenced in 2005.
[6] GORMAN, J., CHUANG, G.-Y., LAI, Y.-T., SHEN, C.-H., BOYINGTON, J. C., DRUZ, A., GENG, H., LOUDER, M. K., MCKEE, K. & RAWI, R. 2020. Structure of Super-Potent Antibody CAP256-VRC26. 25 in Complex with HIV-1 Envelope Reveals a Combined Mode of Trimer-Apex Recognition. Cell Reports, 31, 107488, https://www.cell.com/cell-reports/pdf/S2211-1247(20)30366-1.pdf.
[8] WIBMER, C. K., GORMAN, J., OZOROWSKI, G., BHIMAN, J. N., SHEWARD, D. J., ELLIOTT, D. H., ROUELLE, J., SMIRA, A., JOYCE, M. G., NDABAMBI, N., DRUZ, A., ASOKAN, M., BURTON, D. R., CONNORS, M., ABDOOL KARIM, S. S., MASCOLA, J. R., ROBINSON, J. E., WARD, A. B., WILLIAMSON, C., KWONG, P. D., MORRIS, L. & MOORE, P. L. 2017. Structure and Recognition of a Novel HIV-1 gp120-gp41 Interface Antibody that Caused MPER Exposure through Viral Escape. PLoS Pathog, 13, e1006074. https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1006074.
Investigating lithium-ion battery cathode materials for new generation improvements in sustainable energy solutions
“Ensuring access to affordable, reliable, sustainable and modern energy for all will open a new world of opportunities for billions of people through new economic opportunities and jobs, empowered women, children and youth, better education and health, more sustainable, equitable and inclusive communities, and greater protections from, and resilience to, climate change.”
Lithium-ion batteries are used around the world in everyday portable electronics, in electric vehicles as well as in small power grids. Scientists from the Energy Materials Research Group at the University of the Witwatersrand (Wits), South Africa, study the materials needed to improve the performance, safety, affordability and environmental footprint of lithium-ion batteries in line with important sustainable development goals (SDGs). A range of different cathode materials, as well as battery chemistry is studied to increase the understanding of the materials themselves: how various synthetic routes introduce impurity phases in these materials and how this can be avoided, and what the effects are of intentional doping and co-doping of these materials[2] to explore the impact they have on the structure, performance and impurities formed.
In this work, however, laboratory-based measurements do not often reveal a clear picture of the overall structure of the material, particularly when it contains lower concentrations of impurity phases which are either below the limit at which it can be detected or cannot be resolved from the major phase. It is therefore important to firstly, employ multiple techniques to provide complementary information and secondly, to obtain high-resolution measurements from synchrotron-based techniques which expose far more in terms of other phases in the samples. This is where the GCRF START grant plays a vital part.
“A synchrotron is millions of times more capable than the equipment in our labs in terms of brightness and detail, which makes access through the GCRF START grant to the UK’s national synchrotron, Diamond Light Source (Diamond), so significant.”
GCRF START Co-I, Prof. David Billing[3], Professor in the School of Chemistry and Co-PI of the Energy Materials Research Group at Wits.
“Thus far we have data from various synchrotrons for high resolution X-ray diffraction and total scattering, as well as X-ray absorption spectrometry,” says Group Co-PI, Prof. Caren Billing, lecturer and Associate Professor in the School of Chemistry. “The GCRF START grant provides us with these important experimental opportunities, alongside vital skills training and knowledge exchange for building capacity and training emerging scientists in the energy materials field.”
The ultimate aim of the Group is to address global energy, climate, and health challenges which, amongst other aims, includes enabling better “access to clean and safe cooking fuels and technologies and expanding the use of renewable energy beyond the electricity sector, as well as to increase electrification in sub-Saharan Africa”[4]. To this end, improved battery storage solutions are one of a creative mix of options the Group is examining, with the help of their collaborators, including the GCRF START grant.
“GCRF START asks us to consider questions in our research around the global challenges like: am I using components that are sustainable? Are we using elements which are abundant, affordable, and environmentally compatible?”, explains Prof. Dave Billing. “In doing so, we are mindful that the economic and social situations in Europe are different to Africa: different resources, different engineers, and different environments where the solutions have to work. Everything costs energy, fundamentally, and the whole solution has to fit the community – these are the bigger picture questions that START encourages us to ask.”
Some of the Group’s early career scientists focus on various lithium metal ion phosphate materials, where they are studied with the idea of using earth abundant metal ions of benefit for cost effective production and materials with a possible impact on local mining opportunities[5] to support local economies. Two of these scientists – Michelle Thiebaut from South Africa and Michelle Nyoni from Zimbabwe – are PhD research students at Wits working on lithium iron phosphate and lithium vanadium phosphate respectively, examining them as cathode materials. Both students are supervised and mentored by Prof. Dave Billing and Prof. Caren Billing, and are part of a growing number of female scientists in the Energy Materials Research Group at Wits. Michelle Thiebaut and Michelle Nyoni describe the aims, techniques and motivation for their projects in the case studies below.
Michelle Thiebaut’s research: studying Lithium iron phosphate for battery cathode materials
“When Michelle Thiebaut first started in the group, she referred to herself as “an analogue girl in a digital world”. Michelle is now the forerunner in the group in processing XAS data and has been the only chemistry student to operate the Mössbauer spectroscopy instrument in the School of Physics at Wits.” – Prof. Caren Billing, Prof. Caren Billing, University of the Witwatersrand.
Being a new researcher in a field such as energy materials is both daunting and exciting because this field is always changing and improving. One needs to change and improve one’s skill set just as quickly but the key to continue is finding one’s motivation. My main motivation is seeing how people in South Africa and in other developing countries are struggling with everyday tasks, especially people in the rural areas – tasks like coming home and doing their homework. These tasks are things many people take for granted but I think it is unacceptable that people should be struggling to get by without proper, cheap and a long-lasting access to clean energy and electricity.
My second motivation is our planet. Every person has an obligation to the planet and to live an environmentally cleaner life. By pushing science in the energy materials field means also pushing towards a greener tomorrow. Trying to break through in this field as a young female is still a bit tough with people questioning your skill set and abilities but I do think we owe former female scientists a great deal of respect for paving the way for us.
My research field is energy materials, specifically investigating the cathode material LiFePO4 found in lithium-ion batteries. My focus with this material is to find a low cost, low energy synthetic route and to possibly improve the performance. LiFePO4 is a naturally occurring mineral but can also be synthesised in a lab. This naturally occurring mineral is not phase pure[6], meaning that the iron is commonly mixed with other metals such as manganese, magnesium and calcium which lowers the electrochemical performance[7]. Cathode materials are the positive electrodes of batteries and host the mobile ions (in this case lithium). The mobile ions are the ions that are removed from the structure when the battery is being charged and when the battery discharges (depletes) the ions are inserted back into the structure[8]. Current cathode materials are not only expensive to produce but also have some safety issues like overheating and short-circuiting associated with them – challenges we want to overcome.
Compared with other cathode materials, LiFePO4 has the advantage of being environmentally friendly, meaning there are no toxic materials presents, relatively cost efficient (no expensive metals/rare earth metals needed to synthesise the material) and is structurally and thermally stable. This means that the structure does not collapse with the removal of the mobile ions and the structure prevents the battery from overcharging as well as overheating, making this material safer to use[9]. However, one of the main disadvantages of LiFePO4 is the electrochemical performance such as the ionic (movement of ions through the crystal lattice)[10] and electronic (the ability to conduct or resist electric current), which are both important properties for cathode material. The mobile ions are restricted to movement through a 1-dimensional channel. Overcoming these problems has been the main focus for most research groups[11].
In my research, the electrochemical performance can be improved by doping with a selection of different metal ions. Inserting small amounts of metal ions into the structure can improve the battery performance differently depending on the metal. For example, nickel improves the stability of the structure and enhances the movement of lithium through the structure; copper improves the conductivity and improves the reversibility of the lithium ions in the structure; and manganese improves the reversibility as well as the stability of the structure.
Exploring materials through multiple techniques and collaborative efforts
To fully understand my material, it is very important to understand how the structure changes with small changes in my synthetic method and it is the collaborative effort between the Chemistry and Physics Departments at my university – the University of the Witwatersrand – which makes this possible, and through access to world class synchrotron sources to utilise the benefits of synchrotron data to further characterise my materials. Selected samples were sent to the Synchrotron source at Brookhaven National Laboratory (NSLS-II) in the USA and to Diamond Light Source, the UK’s national synchrotron (Diamond), with access to Diamond provided by the GCRF START grant. We have obtained data from synchrotron X-ray diffraction and total scattering, as well as X-ray absorption spectroscopy. Having remote access to state-of-the-art synchrotron equipment in this COVID-19 travel restricted world is heaven-sent as the research can continue even when no travelling is allowed.
To thoroughly characterise my synthesised materials, I have made use of our lab-based diffractometers in the Chemistry department at Wits as well as the Mössbauer spectrometer and the Raman spectrometer in the Physics department. Mössbauer spectroscopy and Raman spectroscopy are very useful for identifying crystalline (presence of long-range order of the atoms – regular arrangement of atoms over a longer distance) as well as amorphous (only the presence of short-range order of atoms – regular arrangement of atoms but only over a short distance) species in my samples. It is important to identify all the crystalline and amorphous species in the sample as impurities can occur in both forms and could negatively affect the battery performance.
Mössbauer spectroscopy is also useful for identifying the different local iron (Fe) environments present in my sample and to determine the form of iron – (oxidation state – Fe2+ is the desired state in my samples). Raman spectroscopy aids as a structural fingerprint that can be used to determine the identity of one’s material and is also useful in identifying any impurities present that the lab diffractometers could not detect due poor detection limits or due to the phase being amorphous. Synthesised samples can have a mixture of the desired product as well as impurities. There could be multiple sources for the formation of impurities but the most common causes can be either synthesis related (impurities that are formed due to a specific synthetic route) or impurities formed due to sensitivity to air (being exposed to air could cause some small changes like a change in oxidation state of a metal). Impurities can block the channel and subsequently the movement of the mobile ion and negatively affect the performance of the material and the battery.
“Michelle Nyoni is a lady who, against many odds, is striving to obtain her PhD in energy materials. Impacted currently by Covid19 travel restrictions, Michelle is normally an ‘out-of-seat’ student who works full time at Chinhoyi University of Technology (CUT) in Zimbabwe and comes to South Africa for laboratory experiments at the University of the Witwatersrand during her teaching breaks as facilities for her research topic at CUT are limited. She has worked hard during her visits here to gather sufficient data so that she can process it when she returns home and brings with her a great deal of positivity and energy on each visit.” – Prof. Caren Billing, Prof. Caren Billing, University of the Witwatersrand.
While working in the farming sector in my home country of Zimbabwe, I realised that we are blessed with abundant renewable sources of energy – wind and solar – yet hindered by the challenge of how to store this energy effectively. This is where the subject of batteries came into my life and where my current PhD research area fits in. I am investigating lithium vanadium phosphates as cathode materials for lithium-ion batteries. I began my PhD studies part-time, in 2017, at the University of the Witwatersrand in South Africa under the supervision of Prof. Caren Billing and Prof. Dave Billing, while working as a Chemistry Lecturer at CUT in Zimbabwe.
The inspiration for my research is the fact that South Africa is one of the biggest vanadium producers in the world and Zimbabwe is one of the biggest lithium producers in the world. Therefore, if the raw materials are locally available it will hopefully mean reduced cost of battery production. My research is directly linked to Sustainable Development Goal (SDG) 7 concerning affordable and clean energy but by contributing to SDG 7, my research also contributes to achieving SDGs 1-6 and 8-9.
The aim of my research is to do much of the material characterisation by focussing on understanding what is happening at the atomic level, asking questions like: what is happening with the structure and the material? Lithium vanadium phosphate materials have been made but is this synthesis method reproducible? Does it work for upscaling to commercial levels? How do slight changes within the synthesis (preparation method) affect the material? How does adding a dopant manipulate the electrochemical properties of my material?
Lithium vanadium phosphates are potentially effective because their various properties are attractive – they have a high thermodynamic and kinetic stability, and studies have shown they possess the potential to have very good electrochemical properties, which means they will have high specific energy, high working voltage, and good cycle stability. Normally, as batteries get older, the cycling gets poorer and poorer but lithium vanadium phosphate materials have good cycle stability and a lower price tag so they are not as expensive as some of the alternative materials that can be used. There are also other advantages. Lithium vanadium phosphate materials provide improved safety, phosphates are more environmentally friendly than some other materials[12], and the vanadium contributes to the energy density as well as the voltage of the cells – in fact, our Lithium vanadium phosphate materials can reach voltages of over 4 volts! A key application is in electric vehicles which will benefit from increased length of travel due to the cycle stability of Lithium vanadium phosphate materials and the higher specific energy density, amongst other improvements due to the advantages described above.
The cathode material determines the voltage and capacity of a battery and the cathode in a lithium-ion battery is the positive electrode, which is normally a metal oxide that is responsible for being the source of lithium ions that carry the electric current when a battery is in use (discharging)[13]. There are various components that contribute to the cost of the battery with the cathode material within the battery usually one of the biggest costs, along with the separator[14]. The lithium vanadium phosphate materials that I am working with are cathode materials, therefore if we can source these locally within the SADC region of Africa, which includes 15 member states, this would make them a lot more affordable and accessible – which is the goal I am driving at.
Cutting edge techniques to determine material characterisation and impact
The techniques I will use in my PhD studies aim to test the lithium vanadium phosphate materials in depth so that I can contribute to research that is already available to help find viable products that can be used in Africa. Techniques include powder X-ray diffraction for the phase identification and Raman spectroscopy to enable me to determine the structural finger-print to ensure I am making the same product each time so that when I do change a parameter, the resulting effect will be clear. The GCRF START grant enables us to use the Diamond synchrotron for variable temperature experiments. Therefore, I would want to look out for how the material changes when we vary the temperature. I would also use XAFS– X-ray absorption fine structure spectroscopy at the B18 beamline at Diamond to study the changes in the neighbourhoods of particular atoms.
Another technique is transition electron microscopy – the determination of particle size and the distribution of those particles as well as the general morphology of the fine particles within the material. Additionally, I want to use STA – Simultaneous Thermal Analysis – to look at the thermal stability to ask a series of questions: how stable is my material and what happens under temperature changes? Does it break down or decompose? How does this effect the overall electrochemical properties because when we use these batteries they will heat up? What is the impact, for example, if I were to leave my phone device with a battery using these materials in the sun – how would the warmth of the sun affect it? Would the structure and performance be impacted? Therefore, I would do extensive electrochemical testing which includes cyclic voltammetry and electrochemical impedance tests, amongst others, to ensure the batteries with these materials are viable in the varied environmental conditions found across Africa, including very warm environments.
The GCRF START grant: a bridge to sustainable growth and life-changing possibility
“Being part of the GCRF START collaboration has certainly taken our work in energy materials in South Africa to the next level!”
Prof. Caren Billing, University of the Witwatersrand
Many of the Group’s research projects are now at the point where data has been measured and obtained and the next learning curve of how to process the data is underway. Progress has been made, Prof. Caren Billing reports, which, without funding from the GCRF START Grant, would have been an even larger hurdle to overcome.The resultis cutting-edge science and capacity building, knowledge exchange and access to the latest techniques and technology, and a new generation of gifted scientists working towards the shared vision of developing novel, green and affordable energy solutions to inspire life-changing possibility in Africa and beyond.
“This is where the GCRF START grant comes in,” says Prof. David Billing, “it provides that bridge. Yes, there’s a skills gap here in Africa but for me that gap is possibly smaller than others; as long as we are staying current on the XRD side we can transition easily and tackle the more challenging newer techniques – there’s a whole suite of them but that will grow – and START gets us there! This is also what you need to get to the higher impact journals; it also to gets us closer to current answers and future possibilities rather than just ‘the best we can do’ with 30-year-old technology.”
“In terms of energy solutions, take the scenario of load shedding (electricity cuts) which poses a huge challenge across countries in Africa. The thought of being able to go off grid is vital.If you think about a rural village which is cooking using wood or charcoal and lighting in the form of paraffin or candles – this is energy poverty. If you can find a cheap, clean, sustainable source of energy to replace these – that would be life changing!”
GCRF START Co-I, Prof. David Billing, Professor in the School of Chemistry and Co-PI of the Energy Materials Research Group at Wits.
[11] Jingkun Li, Zi-Feng. Past and Present of LiFePO4: From Fundamental Research to Industrial Applications. Chem. Volume 5, Issue 1, 10 January 2019, Pages 3-6 (2019), Elsevier. https://doi.org/10.1016/j.chempr.2018.12.012;
V.S.L. Satyavani,A. Srinivas Kumar,P.S.V. Subba Rao.Methods of synthesis and performance improvement of lithium iron phosphate for high rate Li-ion batteries: A review. Engineering Science and Technology 19, Issue 1, March 2016, Pages 178-188. Elsevier. https://doi.org/10.1016/j.jestch.2015.06.002;
Yi, T., Li, X., Liu, H. et al. Recent developments in the doping and surface modification of LiFePO4 as cathode material for power lithium ion battery. Ionics18, 529–539 (2012). https://doi.org/10.1007/s11581-012-0695-y
[12] Hameed, S.A., Reddy, M.V., Sakar, N., Chowdari, B.V.R. & Vittal, J.J.; Royal Society of Chemistry Advances2015, 5, 60630-60637
[14] See: Figure 3. ‘Total material costs of all 10 considered cell chemistries plus Panasonic NCA Use Case differentiated in combined CAM cost, anode cost, and secondary material costs’ in: Wentker, M.; Greenwood, M.; Leker, J. A Bottom-Up Approach to Lithium-Ion Battery Cost Modeling with a Focus on Cathode Active Materials. Energies2019, 12, 504. https://doi.org/10.3390/en12030504
“The atomic structure of proteins provides an intimate insight into these magnificent macromolecules. This knowledge is crucial to truly understand how they function; whether it is to answer a burning question or to manipulate them – either to enhance if their reactions are desirable, or to inhibit if they are harmful.”
Prof. Dirk Opperman, University of the Free State, South Africa
Our research studies the structures of bacterial and fungal oxidoreductases (enzymes) which are possible drug targets to combat infectious disease. The current focus is fungal drug targets for fungal infectious diseases which can be very serious, especially for immune-compromised patients, such as those who are HIV/AIDS positive, organ transplant receivers, patients undergoing chemotherapy, and many more. This research is performed at the University of the Free State’s (UFS’s) Department of Microbial, Biochemical and Food Biotechnology1 and is led by the two Principal Investigators (PIs) of the Biocatalysis group at UFS, Prof. Dirk Opperman, who is a GCRF START Co-Investigator (Co-I), and Prof. Martie Smit. In our laboratory, we have solved the structures of a number of bacterial and fungal enzymes by X-ray crystallography over the past few years, two of the most recent – solving the structures of fungal cytochrome P450 reductase (Dec 2019) and Baeyer-Villiger monooxygenase (Feb 2020) – were assisted by the GCRF START grant and published in the journals Scientific reports2 and Catalysts3respectively, as described later in this article.
The scale of the Fungal infection and drug resistance challenge
Currently, there are three classes of anti-fungal drugs that are used to combat infectious fungal disease, but there is an increasing number of drug resistant (and even multi-drug resistant) fungi against these drugs meaning that these pathogenic fungi have become or are becoming resistant to the current medication used to treat patients. If no drugs are effective against invasive opportunistic fungi, the prognosis for immune-compromised patients is very poor, and many people will die.
“It is therefore imperative that we search for and develop new antifungal compounds to address the growing challenge [of drug-resistance to opportunistic fungi], which impacts countries across Africa, as well as globally. This is especially urgent if the world is to meet the UN’s Sustainable Development Goals of Health and Wellbeing and Food Security by 2030.”
Dr Carmien Tolmie, University of the Free State, South Africa
Fungal infections are often underreported and because of this the extent of the situation is not fully known. In South Africa, this is of particular concern because of our high incidence of HIV/AIDS. For example, one group4 reported that 90 % of HIV/AIDS positive patients on prolonged treatment contract oropharyngeal candidiasis (also called Thrush), an infection caused by a yeast, which is a type of fungus called Candida (Dos Santos Abrantes, McArthur and Africa 2014). However, this is only one statistic, and the problem is much wider. In another example, Cryptococcal meningitis is a deadly brain infection caused by the soil-dwelling fungus Cryptococcus. Worldwide, nearly 220,000 new cases of cryptococcal meningitis occur each year, resulting in 181,000 deaths, most of which occur in sub-Saharan Africa (CDC, 2020).
Using the powerful beams of Diamond’s synchrotron light to determine protein structures
Our quest to find new drug targets involves examining the chemical processes that happen in the fungal cell in order to keep the cell alive. We choose an enzyme – a special type of protein involved in these processes – which will be a good target for anti-fungal medication. The experiments we do to produce our protein crystals include molecular biology, protein expression, purification and crystallisation. We clone the gene that encodes the enzyme and insert it into a suitable host to produce the protein, such as the bacterium Escherichia coli, by protein expression methods. Escherichia coli is easy to manipulate and inexpensive to culture in large volumes. We then isolate the enzyme by protein purification methods which exploit the physicochemical properties of the enzyme to separate the target from the host proteins, and crystallise it before we examine it by X-ray crystallography.
We use an in-house crystallisation robot at UFS to prepare the crystallisation trials and we regularly collect crystal diffraction data via remote access at the macromolecular beamlines of the UK’s national synchrotron – Diamond Light Source (Diamond). This is a great help as we can control the beamline equipment from our offices, so we don’t incur the expense of travelling to the UK to use synchrotron techniques essential for our research. While we have X-ray diffractometers at the University of the Free State, they are not nearly as powerful as the beamlines of the Diamond synchrotron. Diamond’s beamline hardware has been developed to such an advanced stage that data collection can proceed very rapidly, enabling us to collect data much faster (minutes) than at our home sources (days). This high throughput is essential when searching for and identify tiny molecules that might potentially bind to the protein and possibly act as inhibitors, as large libraries of fragments must be screened.
“The brilliant light generated by Diamond (10 billion times brighter than the sun!) enables us to determine the structure of the proteins to extremely high resolutions, as well as structures from small or weakly diffracting crystals that we cannot study with our own laboratory techniques.”
Dr Carmien Tolmie, University of the Free State, South Africa
We use the protein structure to search for small molecules that will bind to the enzyme and possibly stop it from working (act as inhibitors). In Biocatalysis, knowledge of the protein structure can identify ways in which one can change, or mutate, the enzyme to perform the specific reactions desired. If the protein is a drug target, the structure can be used in Structure-Based Drug Discovery to develop new medications. This process can also be used for other applications like developing new pesticides for agriculture. Therefore, the next step in the research process is to use fragment screening methods to identify lead compounds that can be further developed into inhibitors, thus helping develop a next generation drug. The fragment screening is done in collaboration with Prof. Frank von Delft and the XChem group on the I04-1 beamline at Diamond through the GCRF START grant and will be undertaken once Covid19 travel restrictions are lifted.
Solving the structures of fungal cytochrome P450 reductase and Baeyer-Villiger monooxygenase
Recently, we were able to solve and gain new insights into the structures and mechanisms of the fungal cytochrome P450 reductase (CPR) from Candida tropicalis and the Baeyer-Villiger monooxygenase BVMOAFL210 from Aspergillus flavus, research that was made possible by the GCRF START grant. The results were published in the journals Scientific reports and Catalystsrespectively, and the research on the CPR was done in collaboration with scientists from the University of Cape Town5, who are also part of the START project. The publications were co-authored by START PDRAs Ana Ebrecht (first author on CPR paper) and Rodolpho do Aido Machado (co-author on BVMOAFL210).
The CPR plays a pivotal role in primary and secondary metabolism of different species, from bacteria to animals and plants. In fungi, it supplies electrons to enzymes that are vital for the survival of the organism. The CPR mechanism is complex and involves conformational changes that need to be finely tuned to optimise the process. The structural characterisation of the CPR helps to understand how this process occurs and what are the differences with the human homolog, opening the possibility to use it as a drug target. The structure and mutation data of BVMOAFL210 allowed us to better understand the role of the amino acid at a specific position in the enzyme, in terms of regioselectivity (the position in the substrate where the oxygen atom is inserted) as well as the sulfoxidation (the number of oxygen atoms inserted in a sulfur-containing compound). This residue may be used in future studies for directed evolution experiments to evolve the enzyme to catalyse a desired reaction.
In order to achieve the results, we first needed to produce pure protein. The proteins were crystallised by the vapour-diffusion method with the Douglas Oryx Nano crystallization robot located in our crystallography lab in our department. In these experiments, a library of 384 crystallisation conditions were screened and a few conditions yielded crystals. These crystals were cryo-cooled and shipped at liquid nitrogen conditions in a specialised container to Diamond where we collected data on the macromolecular crystallography beamlines through remote access. We processed the data and solved the structure with programs from the CCP4 suite of macromolecular data processing software. The proteins were characterised further by investigating their kinetic properties with several spectrophotometric assays using a UV/Visible light spectrophotometer.
For BVMOAFL210, we created mutations at a specific position and determined how these mutations alter the biocatalytic profile of the enzyme using whole-cell biotransformation experiments, followed by Gas-Chromatography Mass Spectrometry (GC-MS) analyses. In terms of the fungal cytochrome P450 reductase (CPR), the next steps will be to use the CPR for fragment screening to gain further, more detailed insights. This method uses protein crystals of the target enzyme to identify small molecule fragments that bind to the enzyme. These fragments are then elaborated into larger molecules with higher potency, which will hopefully not only inhibit the specific enzyme, but also the growth of pathogenic fungi.
Benefitting from increased research capacity through the GCRF START grant
The grant has contributed greatly to the research capacity of the Biocatalysis and Structural Biology Research group, making it possible to appoint a START Postdoctoral Research Assistant (PDRA) who focused on structural biology research, the research itself partially funded by START. START also helped to develop the skills of the researchers in the group by funding workshops, as well as workshop attendance, and a research exchange of the START PDRA to the UK in 2019. In addition, START has introduced us to world-class scientists at Diamond and other institutions who we can consult if we need advice on our experiments.
“With this sharing of knowledge, capacity building and cutting-edge research enabled by the GCRF START grant, it is our fervent aim to make a lasting, positive impact in terms of sustainable health, well-being and food security solutions now, and well into the future.”
Dr Carmien Tolmie, University of the Free State, South Africa
Click here to read more about the UN’s Sustainable Development Goals
Acknowledgements
We would like to thank Prof. Trevor Sewell from the START Centre of Excellence at the University of Cape Town’s Aaron Klug Centre for Imaging and Analysis for the pivotal role he has played both in GCRF START and Structural Biology in South Africa. The START Centre of Excellence is a collaborative, shared resource where participants in the START programme can access everything they need to get started with their research, such as advice and the necessary equipment which may not be available in their own laboratories elsewhere. This also includes the technological support and expertise to access the UK’s national synchrotron – Diamond Light Source – through the GCRF START grant (such as support with sample preparation, shipping, and remote access experiments).
[1] Research in the Department falls broadly into three main areas: (i) production of safe and novel food products, (ii) biocatalytic production of chemicals or bioremediation of chemical pollution, and (iii) improvement of human and animal health. Our Biocatalysis Group focuses heavily on biocatalysis which involves the use of one or more enzymes, either as cell-free enzymes or enzymes in whole cells, to convert a substrate into a value-added product. This includes converting alkanes, alcohols, fatty acids or monoterpenes into value added building blocks of pharmaceuticals, bio-plastics, cosmetics, flavours and/or fragrances.
[2] Ebrecht AC, Van der Bergh N, Harrison STL, Smit MS, Sewell BT and DJ Opperman (2019). Biochemical and structural insights into the cytochrome P450 reductase from Candida tropicalis. Scientific Reports 9:20088. doi: 10.1038/s41598-019-56516-6: https://www.nature.com/articles/s41598-019-56516-6
[3] Tolmie C,Do Aido Machado R, Ferroni FM, Smit MS and DJ Opperman (2020). Natural variation in the ‘control loop’ of BVMOAFL210 and its influence on regioselectivity and sulfoxidation. Catalysts 10(3): 339. doi: 10.3390/catal10030339: https://www.mdpi.com/2073-4344/10/3/339
[4] Dos Santos Abrantes PM, McArthur CP, Africa CWJ. Multi-drug resistant oral Candida species isolated from HIV-positive patients in South Africa and Cameroon. Diagn Microbiol Infect Dis 2014;79:222–7.
“Africa is a bank when it comes to diseases. The beauty of the GCRF START grant is that it offers us the opportunity to deal with these challenges in Africa, while enabling international collaboration and access to amazing synchrotron facilities with world class results.”
Dr Ikechukwu Anthony Achilonu, Protein Structure-Function Research Unit (PSFRU), University of the Witwatersrand, South Africa
Neglected Tropical Diseases (NTD’s) and Antimicrobial Resistance (AMR) are major challenges threatening the world’s sustainability and development efforts across the spectrum of the UN’s Sustainable Development Goals, causing millions of deaths each year1. My name is Dr Ikechukwu Achilonu and I work in the PSFRU at the University of the Witwatersrand, in South Africa, where I focus on rational design and discovery of new generation anthelminthic and anti-bacterial drug targets to tackle NTD’s and Nosocomial infections by ESKAPE2 pathogens.
To design inhibitors to serve as potential drugs against these pathogens, high-resolution 3D protein structures are needed to test the ability of certain drugs to interact with and inhibit the properties of the proteins being studied. This is made possible with the GCRF START grant which provides us with access to state-of-the-art synchrotron techniques, equipment and expertise at the UK’s world class national synchrotron, Diamond Light Source (Diamond), enabling the X-ray diffraction from protein crystals generated in our research. The GCRF START grant is currently our only conduit to the UK to enable us to conduct these ultra-high-quality experiments.
My recent research success is solving the Schistosomiasis joponicum Glutathione S-transferase (GST) enzyme has been recently published and is a tribute to the value of collaborating with Diamond and GCRF START. The solved structure is among the crystal structures with the highest resolution in the global protein database for this enzyme with two different ligands. Of the authors, Dr Ramesh Pandian is a GCRF START postdoctoral fellow (Host: Prof. Yasian Sayed), and Dr Sylvia Fanucchi, Prof. Yasien Sayed, Prof. Heine W. Dirr, and I – Dr Ikechukwu Achilonu – are collaborators on the GCRF START grant.
The resulting publication emphasises the need to exploit the unique structural diversity between Schistosoma GST and other human GSTs for a rational approach to design new generation anthelminthics. This research, outlined later in this article, will pave the way for the next exciting step in the structural validation of druggable targets for this debilitating and life-threatening Schistosomiasis (Bilharzia), a NTD which infects large numbers of people across several countries each year4.
The extent of the challenge: NTD’s and ESKAPE pathogens
AMR is described by the World Health Organization5 as “one of the biggest threats to global health, food security, and development today…compromising our ability to treat infectious diseases, as well as undermining many other advances in health and medicine.”
NTD’s are a diverse group of communicable diseases caused by a variety of pathogens prevalent in tropical and subtropical conditions, such as viruses, bacteria, protozoa and parasitic worms (helminths). More than a billion people in over 149 countries are impacted by NTD’s with huge costs to developing economies of billions of US dollars every year. My research on NTD’s focuses on Schistosoma and Wuchereria (Wuchereria is a human parasitic worm (Filariworm) that is the major cause of lymphatic filariasis) – both of which display AMR. Populations living in poverty, without adequate sanitation and in close contact with infectious vectors and domestic animals and livestock, are those worst affected by NTD’s6.
The six nosocomial ESKAPE pathogens that exhibit multidrug resistance and virulence are often acquired in health care settings like hospitals (Health Care Acquired Infection/HAI) and feature on the World Health Organization’s ‘Priority Pathogens list’7. The persistent use of antibiotics has provoked the emergence of multidrug resistant (MDR) and extensively drug resistant (XDR) bacteria, which can make even the most effective drugs ineffective. These bacteria have built-in abilities to find new ways to become resistant and can pass on genetic materials which enable other bacteria to become drug resistant. Like NTD’s, this situation is made worse by limited or non-existent infection prevention and control (IPC) programmes, combined with an inadequate water supply, poor sanitation, and a weak hygiene infrastructure in health facilities, which make the burden of HAI several fold higher in low- and middle-income countries than in high-income ones.
Over the past three years, I have been working on the Schistosoma GST enzyme, which is a protein that is involved in detoxification of foreign molecules in the Schistosome parasite that causes the disease Schistosomiasis/Bilharzia. This enzyme very critical to the survival of the parasite. If the enzyme is not functioning properly, the parasite cannot move from egg stage to lava stage. Therefore, if the enzyme could be effectively inhibited, one would have a drug against the parasite.
In my research, I used a molecule which is a natural product found in pomegranate juice called Ellagic Acid. One of my MSc. students, Ms Blessing Akumadu, carried out the study I had conceptualised, showing that this molecule can inhibit the activity of the Schistosoma GST by 66%. The concept was based on an earlier study that ellagic acid and its derivative inhibits Plasmodium GST (duly acknowledged in the forthcoming paper).To be more confident about making extrapolations from the structure of the Schistosoma japonicum GST enzyme, we needed to see that our wet lab results had meaning in 3D high resolution. Using the I038 and I04 beamlines at Diamond, we were able to solve the structure of the 26 kilodalton GST at 1.53 Å resolution and were amazed when we saw the results! We had achieved the highest resolution of any of the several variants of the Schistosoma japonicum GST in the global protein database!
What we observed is the Schistosoma 26 kilodalton GST enzyme (Sj26GST), which is a dimeric protein (it has two similar subunits joined together) with an interface where the subunits are joined. The unique thing about this protein is that the dimer interface can be exploited for drug interaction, unlike other GST’s in humans. The results have enabled us to see where the Ellagic Acid molecule (the potential drug) can bind to the protein. In this parasite we only have two classes of GSTs (in humans there are several) – the 26 kilodalton (kDa) GST and the 28 kDa GST. Having studied the 26 kDa GST, we are now currently working on the 28 kDa GST across the three species of the Schistosoma parasite that infect humans (mansoni, haematobium and japonicum) to see if the Ellagic Acid molecule can also inhibit the 28 kDa GST of the Schistosoma parasite.
The implications of these results mean we can go on and redesign this potential drug molecule because we can see the type of interaction this drug is making with the protein in the 3D images. The more the drug interacts, the stronger the binding. Therefore, we can start to redesign this drug for better binding so that the binding and the efficiency is improved beyond the 66% to maybe 75% or even to 99 % in order to completely shut down the activity of the enzyme at a very low amounts of the drug. Without the high-resolution 3D crystal structure solved using Diamond, we could not do this. We can now exploit this interaction by adding a bond here or a functional group there to improve the strength of the interaction and potency of the drug, and produce new high resolution structures until we have a drug that shuts down the enzyme and produces a new drug target to which the Schistosoma parasite is not resistant to.
The benefit of studying multiple NTD and ESKAPE pathogens simultaneously – the case of nicotinamide adenine transferase enzyme in klebsiella pneumonia and Enterococcus faecium.
In my experience, the techniques one applies to studying one protein can also be applied to the others, which is why I study a range of druggable proteins from several human NTD and ESKAPE pathogens simultaneously. For example, my students and I are examining the nicotinate-nucleotide adenylyltransferaseenzyme which is important in the redox metabolism in both klebsiella pneumonia and Enterococcus faecium. Klebsiella pneumonia is a Gram-negative bacterium resistant to multiple drugs and increasingly resistant to most available antibiotics (it has a thin cell wall surrounded by a protective cell membrane which makes it harder for antibodies to penetrate). In health care settings, patients who require devices like ventilators or intravenous catheters, and patients who are taking long courses of certain antibiotics are most at risk for Klebsiella infections.
The Gram-positive Enterococcus faecium is a human pathogen that causes nosocomial bacteremia, surgical wound infection, endocarditis, and urinary tract infections (it has thick cell walls but no outer membrane). Studying both simultaneously has led to us noticing that the Gram-negative function of the nicotinamide adenine transferase enzyme performs differently from the Gram-positive function of the same enzyme which is exciting because this enables us to infer structural and functional differences between this enzymes from different bacteria that are supposed to perform similar activities in these bacterial pathogens. This has serious implication in the design of a broad-spectrum antibiotics that will target this enzyme in different organisms.
The process of solving protein structures –systematic integration of multi-facted ‘new generation’ theoretical and empirical studies
The approaches we use in the research outlined above are multi-faceted and multi-disciplinary involving the integration of sophisticated ‘new generation’ theoretical and empirical studies and, as already indicated, a range of cutting-edge equipment. To design drugs against disease pathogens, the three-dimensional architecture of the druggable protein is needed. Druggable proteins are proteins that are key to the survival of the pathogen which enables it to replicate inside the human host, either intracellularly or extracellularly. In the case of bacteria, for example, they need to manufacture their own energy and without the enzyme (protein) that makes this possible, the bacteria would not survive. Therefore, if we know the protein’s DNA sequence, we should be able to predict that we can synthesis this protein and develop the druggable target.
To elucidate the crystal structure of these druggable proteins it is possible to use home X-ray sources. Ultimately, however, there is a major advantage of using a powerful synchrotron light source like Diamond over most home X-ray sources because the amount of inferences made from a protein crystal with resolutions in the range of 1.3 – 1.7 Å is unprecedented. Such crystals can be further studied using computational algorithms and can also serve as models for replacement when solving the crystal structure of a protein that is very similar in structure.
In order to generate the recombinant proteins that are druggable targets, we need large amounts of these proteins. For this we use E. coli bacteria like a factory to synthesize our target proteins and carry out a rigorous ‘quality assurance’ process to ensure the proteins we make are pure, stable and homogenous for successful crystallisation experiments. If the protein is an enzyme, it should be able to catalyze the same function it does naturally in an organism (in vivo), inside a test tube (in vitro), which means that whatever we send off to Diamond for X-ray diffraction experiments, we need to be sure that the biological function is intact. To do this, the proteins are assayed (measured) for catalytic activity in vitro (if they are enzymes) and are further characterised using several biophysical techniques to test their structure and the function to the point we are satisfied that the proteins are active, properly folded and can perform the function they are supposed to perform properly (biological activity is maintained).
We purify the proteins to homogeneity using an ensemble of protein chromatography techniques. For ‘quality assurance’, we check the secondary structure content (Far-UV circular dichroism), the tertiary structure (intrinsic and extrinsic fluorescence spectroscopy); and the quaternary structure (using size exclusion HPLC). Every physical interaction between two biomolecules generates heat or absorbs heat as part of the function of the protein to interact with another molecule (molecular recognition). Therefore, to estimate the thermodynamics of protein-ligand interaction (interaction ability), we use isothermal titration calorimetry.
Once we are satisfied that the proteins are of good quality and quantity, we begin protein crystallisation. I like to use the analogy that structural biologists are akin to photographers and using X-ray diffraction is like taking a snapshot of the protein which has been forced to assemble as repeating units in a crystalline form. There are several approaches we use. We can either conduct crystallisation manually, or we can do it robotically using our in-house Douglas Instrument Oryx 8 crystallography robot for high throughput protein crystallisation.
To solve the protein structure – sometimes in complex with potential drugs – X-ray light is invaluable. However, X-rays have very short wavelengths and if you keep bombarding a protein crystal with X-rays, it will heat it up and gradually fall apart because of X-ray damage. By changing the protein from solution to a crystal, and then embedding it in a liquid able to resist the intense bombardment of X-ray (cryo-protection) by freezing it at the temperature of liquid nitrogen (-196 degrees Celsius), the sample stays stable and the X-rays will last 100x longer in the X-ray beam! Our crystal samples are therefore sent to Diamond in special containers which keep the samples crystals frozen. GCRF START makes it very easy to ship our crystals to the UK without lots of red tape. We use DHL and we have an easy system which means it only takes a few days to get to Diamond ready for the experiments; the GCRF START grant helps this process at no cost to us.
We can use an X-ray diffractometer such as a small in-house diffractometer to solve the protein structure, or the ultimate technique for X-ray crystallography – a massive synchrotron like Diamond! Diamond’s I03 and I04 beamlines are used to fire X-rays onto our crystal samples. The X-ray will be scattered based on the structure of the protein/molecule (X-ray diffraction). Then, using mathematical modelling and other techniques, the 3D structure of the protein is resolved, either on its own or in a complex with a drug or other biomolecules such as DNA.
My theoretical (computational) studies involve several molecular modelling techniques to model the atomistic behaviour of our proteins of interest, especially how the dynamics of these proteins are altered upon drug binding. We also screen various in silico (virtual) libraries (including PubChem, ZINC and Drug Bank databases) for potential hits that we further extend to the theoretical studies, including high-throughput virtual screening, induced-fit ligand docking, molecular dynamic simulation and free binding energy calculations.
Sometimes, however, the process provides a few surprises, which was the case in our journey to solving the structure of the Schistosoma japonicum GST enzyme. We usually send our crystals to Diamond at the same time as the other groups in South Africa because we have the same slots for data collection. Two weeks before we were supposed to ship our crystals, we found that none were ready yet to ship. This wasn’t good. Therefore, to hurry things along, I asked one of my MSc students, Jessica Olfsen, who knows the crystallization robot very well to generate the crystals we needed within one week. This she did, but they were very ugly looking crystals! I did not think we would get any data out of these crystals but at least we had crystals to send. I so doubted their usefulness that when I collected the data, I didn’t even want to look at it, let alone solve the protein structure. In my disappointment I asked one of our new post-docs (Dr Pandian) who needed a task to look at it. The resulting 3D high resolution structure blew him away! When we both looked at it, we saw just how beautiful it was. I had written the paper already, but it had needed more experimental data for publishing so now I had it! The structure was solved at unprecedented resolution and the paper was accepted for publishing.
As a structural biologist, drug discovery and protein biochemistry are completely intertwined and multidisciplinary in the sense that I cannot do without a synthetic chemist (organic chemist). These are the people who will synthesize the drug. For example, once we add the extra functional groups, it will improve the binding capacity of the drug. The synthetic chemist will then go into the laboratory, resynthesize the drug before I will return to my lab and see if the inhibition improves. When an improvement in the inhibition is apparent, we go through the whole process again: recrystallize the protein with the proposed drug, and then use Diamond again to produce another high-resolution 3D structure of the re-modelled drug molecule and protein interaction.
“This whole drug discovery process can take some time, so it is vital that we have access to world class synchrotron facility like Diamond for the lifetime of this research we until we have the results needed to tackle the NTD’s and ESKAPE pathogens which impact countless lives.”
Dr Ikechukwu Anthony Achilonu, Protein Structure-Function Research Unit (PSFRU), University of the Witwatersrand, South Africa
Read more here about the UN’s Sustainable Development Goals.
More about Dr Achilonu
Dr Ikechukwu Anthony Achilonu is Senior Researcher and the Interim South African Research Chair (NRF/SARChI) in Protein Biochemistry and Structural Biology at the Protein Structure-Function Research Unit (PSFRU), School of Molecular and Cell Biology, Faculty of Science, University of the Witwatersrand in South Africa.
[1] Transforming our world: the 2030 Agenda for Sustainable Development – A/RES/70/1. New York, USA: United Nations; 2015.
[2] ESKAPE is an acronym for the group of bacteria, encompassing both Gram-positive and Gram-negative species, made up of Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species.
[3] Authors: Blessing O. Akumadu, Ramesh Pandian, Jessica Olfsen, Roland Worth, Monare Thulo, Tshireletso Mentor, Sylvia Fanucchi, Yasien Sayed, Heini W. Dirr, Ikechukwu Achilonu. Title: Molecular basis of inhibition of Schistosoma japonicum glutathione transferase by ellagic acid: insights into biophysical and structural studies. Publication: Molecular and Biochemical Parasitology (2020);Elsevier. https://doi.org/10.1016/j.molbiopara.2020.111319
