Center for Tropical and Emerging Global Diseases
University of Georgia
Vasant Muralidharan, associate professor in Franklin College‘s Department of Cellular Biology and member of CTEGD, is the featured guest in this episode of People, Parasites, and Plagues. He discusses his work with the Plasmodium parasite and his personal experience with malaria.
by Donna Huber
The parasite that causes malaria was discovered more than 125 years ago, but much is still unknown about this complex, single-celled organism. Researchers in the University of Georgia’s Center for Tropical and Emerging Global Diseases, however, have uncovered the role of one of the parasite’s essential proteins, offering new insights for vaccine and drug development.
Plasmodium falciparum causes the deadliest form of malaria, a disease the World Health Organization estimates killed more than 600,000 people worldwide died in 2022. A large majority of those deaths were children under the age of 5.
Historically, the parasite has been difficult to study due to its complex lifecycle, which includes three stages. One occurs in the mosquito, while the liver and blood stages take place in humans. The blood stage is when the infected person exhibits symptoms of malaria.
In the blood stage, the parasite invades red blood cells (RBCs) where they replicate and can be transmitted to the mosquito. The receptor-ligand complexes that enable RBC invasion have been well-studied and it is one of the targets of anti-malarial vaccines currently in clinical trials. But questions still remain.
“How does the parasite know it has encountered a red blood cell?” asked Vasant Muralidharan, associate professor in Franklin College’s Department of Cellular Biology and leader of the Muralidharan Research Group, where the study took place.
Interested, the team took a closer look at a protein called RON11, which is sent to a pair of unique club-shaped secretory organelles known as the rhoptry (Greek for club) that houses proteins needed to invade the RBC.
“When we knocked out this protein, we found that the parasite could do everything it usually does – create a putative pore in the membrane of the RBC, send proteins needed for parasite invasion through this putative opening into the RBC – but the parasite itself cannot enter the red blood cell,” Muralidharan explained. “If a parasite cannot enter the red blood cell, the life cycle is interrupted and the parasite dies.”
And then things got really interesting.
“We found that the parasites lacking RON11 were only producing half the rhoptry proteins, which are used in invasion,” Muralidharan said.
While it is known that Plasmodium parasites have two rhoptry organelles, they are so teeny-tiny they have been relatively understudied due to a lack of proper tools. However, new tools and techniques are emerging. David Anaguano, a cellular biology graduate student who led the study, traveled on a Daniel G. Colley Training in Parasitology fellowship to the Absalon Laboratory at Indiana University School of Medicine to learn a new tool known as Ultrastructure Expansion Microscopy.
“Electron microscopy is labor intensive, and since it uses thin slices of the parasite you are never sure if what you’re looking for really isn’t there or just not in the slice of the sample you have,” Muralidharan said. “Expansion microscopy is like using light microscopy but with a special gel to expand the cell proportionately in all directions. Thus, you don’t get the distortion you would with just an enlarged cell and you can image the entire infected cell in all dimensions. It has been a real game changer.”
As reported in the PLoS Biology paper, the Muralidharan group generated for the first time a Plasmodium cell with only one rhoptry organelle when they removed RON11 from malaria parasites.
“It’s not unusual for an organism to have a backup copy, but we can see that the parasite can create the first rhoptry just fine – without defect – but the second one that should form during the end of the replication cycle never forms,” Muralidharan said. “Why is that?”
As it appears that this second rhoptry is needed for RBC invasion, understanding the mechanisms that control its development could open up new targets for vaccine and drug treatment discovery as well as answering crucial questions like whether the two rhoptries are identical.
“This has been a long unanswered question,” Muralidharan said. “Now with this RON11 knockout parasite that doesn’t form a second rhoptry, we have the tools to answer it.”
Malaria is a global and deadly human disease caused by the apicomplexan parasites of the genus Plasmodium. Parasite proliferation within human red blood cells (RBCs) is associated with the clinical manifestations of the disease. This asexual expansion within human RBCs begins with the invasion of RBCs by P. falciparum, which is mediated by the secretion of effectors from 2 specialized club-shaped secretory organelles in merozoite-stage parasites known as rhoptries. We investigated the function of the Rhoptry Neck Protein 11 (RON11), which contains 7 transmembrane domains and calcium-binding EF-hand domains. We generated conditional mutants of the P. falciparum RON11. Knockdown of RON11 inhibits parasite growth by preventing merozoite invasion. The loss of RON11 did not lead to any defects in processing of rhoptry proteins but instead led to a decrease in the amount of rhoptry proteins. We utilized ultrastructure expansion microscopy (U-ExM) to determine the effect of RON11 knockdown on rhoptry biogenesis. Surprisingly, in the absence of RON11, fully developed merozoites had only 1 rhoptry each. The single rhoptry in RON11-deficient merozoites were morphologically typical with a bulb and a neck oriented into the apical polar ring. Moreover, rhoptry proteins are trafficked accurately to the single rhoptry in RON11-deficient parasites. These data show that in the absence of RON11, the first rhoptry is generated during schizogony but upon the start of cytokinesis, the second rhoptry never forms. Interestingly, these single-rhoptry merozoites were able to attach to host RBCs but are unable to invade RBCs. Instead, RON11-deficient merozoites continue to engage with RBC for prolonged periods eventually resulting in echinocytosis, a result of secreting the contents from the single rhoptry into the RBC. Together, our data show that RON11 triggers the de novo biogenesis of the second rhoptry and functions in RBC invasion.
David Anaguano, Opeoluwa Adewale-Fasoro, Grace W Vick, Sean Yanik, James Blauwkamp, Manuel A Fierro, Sabrina Absalon, Prakash Srinivasan, Vasant Muralidharan. PLoS Biol. 2024 Sep 18;22(9):e3002801. doi: 10.1371/journal.pbio.3002801. eCollection 2024 Sep.
My name is Grace Vick and I am a 4th year infectious diseases PhD candidate in Vasant Muralidharan’s lab. I’m originally from North Carolina and received my Bachelor’s of Science in Biology from Western Carolina University. After graduating undergraduate, I completed an internship at the Defense Forensic Science Center doing forensic biology research. After that, I spent 2 years as an ORISE Fellow at the Centers for Disease Control and Prevention, studying and identifying genetic markers of multi-drug resistant strains of Neisseria gonorrhoeae. I came straight to UGA through the ILS program after my fellowship at CDC.
What made you want to study science?
Ever since I was little, I’ve always spent a lot of time being outside in nature and enjoyed figuring out the intricacies of how things work. During my undergraduate, I was able to explore the different areas of science and found the molecular biology of genetics to be an interesting field that is highly translatable and still vastly unknown. After I spent a few years gaining lab experience and an appreciation for the public health concerns of infectious diseases at the CDC, I knew I wanted to pursue a PhD in that field which brought me to UGA.
Why did you choose UGA?
My experience at the CDC offered the opportunity to learn about diseases and public health issues across all sectors and countries, which led me to learn more about parasitic diseases. Previously, I knew nothing about these diseases but as I learned more about their complex and fascinating life cycles and how these diseases of poverty impact people around the world, I was captivated by this research. Because I was really interested in spending my PhD studying infectious and parasitic diseases, I found out about the CTEGD at UGA and that is what brought me here. The CTEGD is a really wonderful environment for trainees to be exposed to exciting and diverse parasitology research, and I’ve really enjoyed my experience here.
What is your research focus and why did you choose it?
Our lab works on the deadliest form of malaria, Plasmodium falciparum. P. falciparum kills over half a million people each year, with the majority of those deaths being children under the age of 5. Our lab is interested in understanding the molecular mechanisms that are essential to asexual blood stage of this parasite. My work specifically focuses on determining the role of previously unknown proteins that we have discovered are essential for asexual stage invasion of merozoites into host red blood cells. Using a combination of genetic engineering, molecular, and cellular biology techniques, I aim to determine the molecular function of these proteins in the human asexual stage invasion of red blood cells.
Have you received any awards or honors?
In addition to receiving the NIH T32 Predoctoral Fellowship, I have been invited to present at multiple national and international conferences such as Molecular Parasitology Meeting in Massachusetts and Molecular Approaches in Malaria in Lorne, Australia where I won a poster award.
What are your career goals?
When I graduate with my doctoral degree, I hope to either join governmental research or the industry sector. If I decided to head into governmental work, I would choose a career at the CDC where I could continue working in the parasitology research field and apply current public health policies to the international parasitology field. If I decide to join the biomedical industry sector, I would want to work in Research and Design at a company that designs therapeutics and diagnostics for disease prevention and treatment.
What do you hope to do for your capstone experience?
I would really love to experience fieldwork in a malaria-endemic region. I think having the experience of meeting people and learning firsthand how this disease affects millions of people every day would be very eye-opening for me since I have only seen the lab side of malaria. The ability to experience fieldwork would give me a broader experience with how malaria is researched and treated outside of the lab environment and in rural lab environments. I would love to visit Africa or South East Asia to conduct fieldwork in a malaria-endemic environment.
What is your favorite thing about Athens?
Obviously, I love the food in Athens! I love going downtown to grab food and drinks on the weekend. Otherwise, I enjoy getting out and exploring the green spaces and parks that Athens has to offer such as Sandy Creek and the North Oconee Greenway with my husband and dog.
Any advice for a student interested in this field?
I would say the best advice is to read and soak up as much as you can about parasitology both before you get into the field and after. A lot of research has overlap between different parasites and it’s helpful to know about other parasitic diseases that might not be your main focus. Plus, parasites are fun! 🙂 My other advice in general for starting graduate school is to always reach out to students in labs you’re interested in joining. Students are pretty much always willing to help give clear insight into lab dynamics, mentorship of the PI, and generally how life working in that lab is. That information is all really helpful to know when choosing which lab to join!
Support trainees like Grace by giving today to the Center for Tropical & Emerging Global Diseases.
Apicomplexan parasites exhibit tremendous diversity in much of their fundamental cell biology, but study of these organisms using light microscopy is often hindered by their small size. Ultrastructural expansion microscopy (U-ExM) is a microscopy preparation method that physically expands the sample by ~4.5×. Here, we apply U-ExM to the human malaria parasite Plasmodium falciparum during the asexual blood stage of its lifecycle to understand how this parasite is organized in three dimensions. Using a combination of dye-conjugated reagents and immunostaining, we have cataloged 13 different P. falciparum structures or organelles across the intraerythrocytic development of this parasite and made multiple observations about fundamental parasite cell biology. We describe that the outer centriolar plaque and its associated proteins anchor the nucleus to the parasite plasma membrane during mitosis. Furthermore, the rhoptries, Golgi, basal complex, and inner membrane complex, which form around this anchoring site while nuclei are still dividing, are concurrently segregated and maintain an association to the outer centriolar plaque until the start of segmentation. We also show that the mitochondrion and apicoplast undergo sequential fission events while maintaining an association with the outer centriolar plaque during cytokinesis. Collectively, this study represents the most detailed ultrastructural analysis of P. falciparum during its intraerythrocytic development to date and sheds light on multiple poorly understood aspects of its organelle biogenesis and fundamental cell biology.
Benjamin Liffner, Ana Karla Cepeda Diaz, James Blauwkamp, David Anaguano, Sonja Frolich, Vasant Muralidharan, Danny W Wilson, Jeffrey D Dvorin, Sabrina Absalon. Elife. 2023 Dec 18:12:RP88088. doi: 10.7554/eLife.88088.
The malaria-causing parasite, Plasmodium falciparum completely remodels its host red blood cell (RBC) through the export of several hundred parasite proteins, including transmembrane proteins, across multiple membranes to the RBC. However, the process by which these exported membrane proteins are extracted from the parasite plasma membrane for export remains unknown. To address this question, we fused the exported membrane protein, skeleton binding protein 1 (SBP1), with TurboID, a rapid, efficient, and promiscuous biotin ligase (SBP1TbID). Using time-resolved, proximity biotinylation, and label-free quantitative proteomics, we identified two groups of SBP1TbID interactors: early interactors (pre-export) and late interactors (post-export). Notably, two promising membrane-associated proteins were identified as pre-export interactors, one of which possesses a predicted translocon domain, that could facilitate the export of membrane proteins. Further investigation using conditional mutants of these candidate proteins showed that these proteins were essential for asexual growth and localize to the host-parasite interface during early stages of the intraerythrocytic cycle. These data suggest that they may play a role in ushering membrane proteins from the PPM for export to the host RBC.
David Anaguano, Watcharatip Dedkhad, Carrie F Brooks, David W Cobb, Vasant Muralidharan. J Cell Sci. 2023 Sep 29;jcs.260506. doi: 10.1242/jcs.260506
It’s a microscopic parasite, invisible to the naked eye but common in tropical and subtropical regions throughout the world. Each year, millions of people are infected by Plasmodium and exposed to an even more debilitating—and often deadly—disease: malaria.
Malaria is one of the deadliest diseases known to man. It can lead to extreme illness, marked by fever, chills, headaches and fatigue. More than half the world’s population is at risk of contracting the disease, and those who develop relapsing infections suffer a host of associated costs.
Limited educational opportunities and wage loss lead to an often unbreakable cycle of poverty. Vulnerable populations are most at risk.
“When I’m teaching in an endemic area like Africa, it isn’t unusual to find a student who needs to sleep during part of the workshop because they have malaria,” researcher Jessica Kissinger said.
It’s a challenge she and her collaborators in the University of Georgia’s Center for Tropical and Emerging Global Diseases (CTEGD) are trying to combat.
When the Center was established in 1998, there were only a couple of faculty members studying Plasmodium. Now, 25 years later, it has become a world-class powerhouse of multidisciplinary malaria research. Scientists examine various species of the dangerous parasite, studying its life cycle and the mosquito that transmits it.
While Plasmodium seems to have superpowers that allow it to evade detection and resist treatment, CTEGD researchers are working together to innovate and transfer science from the lab to interventions on the ground.
Plasmodium is a complex organism, and studying it is like putting together a jigsaw puzzle. Some researchers contribute pieces related to the blood or liver stages of the parasite’s lifecycle, while others provide insights about hosts interactions. One way UGA’s research connects with the global effort to eradicate malaria is PlasmoDb—a resource derived in part from Kissinger’s research that is now part of a host of databases under the umbrella of The Eukaryotic Pathogen, Vector and Host information Resource (VEuPathDB).
“Our group has been able to help many others when their research question crosses into an –omic,” Kissinger said, referring to in-house shorthand for domains like genomics, proteomics and metabolomics.
Kissinger, Distinguished Research Professor of genetics in the Franklin College of Arts & Sciences, became interested in malaria and Plasmodium during her postdoctoral training at the National Institutes of Health (NIH). Working from an evolutionary biology perspective, she’s interested in how the parasite has changed over time.
“I see it as an arms race,” Kissinger said. “I want to understand what moves they have and can make.”
To understand the parasite, you must dive deep into its genetic code.
Kissinger paired her work in Plasmodium genomics with her interest in computing by helping create the database with information from the Plasmodium genome project completed in 2002. The Malaria Host-Pathogen Interaction Center, one of her projects at UGA, was a seven-year, multi-institutional effort funded, in part, by NIH to create data sets that could be used in systems biology of the host-pathogen interaction during the development of disease.
“Wouldn’t it be neat if, from the beginning of infection all the way to cure, you knew everything that was going on in the organism all the time?” Kissinger said, noting the project’s goal.
They generated terabytes of data that, along with data from the global research community, are publicly accessible and reusable through PlasmoDB and other resources.
Being part of a group that is studying so many different aspects of malaria helps put Kissinger’s research into perspective. Now, in addition to understanding the parasite, she also thinks about tools needed to facilitate research from peers.
David Peterson, professor of infectious diseases in the College of Veterinary Medicine, noted that low-tech solutions have mitigated malaria’s human costs. He acknowledged, however, that their long-term goals required more.
“We have to acknowledge that low-tech solutions, such as mosquito nets, have saved lives,” Peterson said. “But to develop the high-tech solutions that will one day end malaria, we need basic research.”
Pregnant women are particularly vulnerable to malaria because their existing immunity to malaria fails to protect them during pregnancy. Placental malaria often results in premature birth and low birth weight.
Peterson is interested in a binding protein that allows the parasite to adhere to the placenta. While many P. falciparum parasites have only one gene copy that encodes the placental binding protein, Peterson is investigating Plasmodiumisolates with two or more slightly different copies.
But why isn’t one copy enough?
That is the primary question Peterson is focused on. He wants to understand how Plasmodium uses extra copies to evade the immune system, distinguishing the role of each requires tools that Vasant Muralidharan, associate professor of cellular biology, has.
Muralidharan’s interest began when he contracted malaria himself. Through access to good health care, he made a full recovery, but the pain he endured remained. He wanted to understand this parasite. Even more, he wanted to make an impact with research.
His graduate training focused on biophysics, but soon his interest in Plasmodium resurfaced. He discovered there was a lack of tools to study the parasite on a genetic level.
“It’s like a house of cards, and each card is a gene,” Muralidharan said. “You can remove one and see what happens—does the house fall or remain standing?”
In the days before CRISPR/Cas9, there wasn’t a precise way to remove genes. Muralidharan is among the pioneers of gene-editing techniques in Plasmodium.
Like Peterson, Muralidharan focuses on proteins secreted by the parasite. He studies the largely unknown process that allows the parasite to invade a red blood cell (RBC), replicate and escape. The lack of tools was a major hindrance, so Muralidharan created new ones.
These tools have been used by Muralidharan’s CTEGD and CDC colleagues to see how drugs might fail. Muralidharan’s laboratory can create mutant Plasmodium parasites that become resistant to a particular drug, and genome sequence databases allow researchers to check if that mutant is already circulating in malaria endemic regions.
Plasmodium vivax is the predominant malaria parasite in Southeast Asia. It causes “relapsing malaria” during which some parasites go “dormant” after entering the liver instead of reproducing. This phase is a major obstacle for current treatments.
CTEGD Director Dennis Kyle, GRA Eminent Scholar Chair in Antiparasitic Drug Discovery and head of the Department of Cellular Biology, became fascinated with the Plasmodium parasite early in his career, spending time living in Thailand and working in refugee camps where malaria is prevalent.
“When I first got to the refugee camp and saw the situation people were living in, I questioned my decision to become a scientist in the lab instead of becoming a physician,” Kyle said, recalling a camp he worked in that housed about 1,300 kids between the ages of 2 and 15. “There was a guy who was a leader in the group who probably had no more than an early high school education. He said, ‘Look at what you can do—you might generate something that would benefit all of us. The physicians we have in the camp can only work on a few people at a time.’”
Kyle’s laboratory is looking to repurpose medications that have antimalarial properties, a safe way to reduce the development time from lab to clinical use. He’s optimistic we will see a drug treatment that eliminates vivax malaria.
“That’s where UGA is playing a major role,” he said. “The Gates Foundation funded us to develop tools to study the dormant parasite in the liver. And we’ve been successful.”
One of Kyle’s collaborators is Samarchith Kurup, assistant professor of cellular biology, who studies the human immune response to Plasmodium infection.
“We use mouse models to delve into the fundamental host-parasite interactions, which you cannot do practicallyin humans,” Kurup said. “Our understanding of these fundamental processes gives rise to newer and better vaccination approaches and drugs.”
Another important CTEGD addition is Chet Joyner, assistant professor of infectious diseases, whose work has helped make it easier to study dormant parasites stateside.
Like other Plasmodium researchers, Joyner became interested in parasites at an early age. During an undergraduate parasitology class, he discovered how little was known about P. vivax. He was already interested in how diseases develop, so for graduate school he focused on the liver stage of vivax malaria. However, it was a difficult task.
“At the time, the technologies weren’t there,” Joyner said. “Dennis was working on his system, but it wasn’t on the scene yet. I changed from studying the parasite to studying the animal model to understand pathogenesis and immunology in humans.”
Joyner joined UGA after completing his postdoctoral training at Emory University, where he developed a non-mouse animal model to study vivax malaria.
“We have to go to [Thailand] where people are infected and collect blood samples and then feed mosquitoes these samples to do the necessary studies,” Kyle said. “That’s been very impactful. We’ve gotten a lot of data out of it, and now with Chet’s model it all can be done under one roof.”
Joyner wants to understand the human immune response with a focus on vaccine development. Building on Muralidharan’s and other researchers’ findings of how the parasite interacts with the RBCs, Joyner’s vaccine program targets a specific protein in the parasite that inhibits the development of immunity.
“My colleagues have shown that if you knock this protein out in the parasite, the immune response in mice is actually great, and we are now working together to evaluate this in non-mouse models.” Joyner said.
Joyner also has collaborated with Belen Cassera, professor of biochemistry, to screen drug compounds. Cassera’s training focused on metabolism to find drug targets. She is particularly interested in how a drug functions.
“If we understand how the drug works, it will help us predict potential side effects in humans,” Cassera said. “We can’t predict everything, but knowing how it works gives you some confidence in whether it will work in humans.”
Cassera is focused on finding drugs that will treat the more lethal Plasmodium falciparum, the predominant species in Africa, which is rapidly becoming resistant to current treatments. Her work is complementary to Kyle’s.
“They run certain assays for the liver-stage infection, and our lab benefits because we want to know if the drug we are developing is specific for the blood stage or can tackle all stages,” Cassera said.
“Malaria is a vector-borne disease transmitted by a mosquito. You need to tackle not only the parasite in the human but also stop its transmission,” Cassera said. “CTEGD is unique because we can study the whole life cycle, including the mosquito.”
Michael Strand, H.M. Pulliam Chair of Entomology in the College of Agricultural and Environmental Sciences and a National Academy of Sciences Fellow, is an expert on parasite-host interactions. Instead of the human host, he is interested in mosquitoes. Recent work indicates blood feeding behavior of mosquitoes strongly affects malaria parasite development while the gut microbiota of mosquitos could lead to new ways to control populations. Having the SporoCore insectory on campus aids his research.
Established in 2020, SporoCore, under the management of Ash Pathak, assistant research scientist in the Department of Infectious Diseases, provides both uninfected and Plasmodium-infected Anopheles stephensi mosquitoes to researchers at UGA and other institutions. Like Joyner’s animal model, the insectory allows for research to be done in the U.S. that would otherwise require field work in an endemic country.
Old-school interventions like mosquito nets, combined with new drug therapies, have reduced the number of malaria deaths, which declined over the last 30 years before rising slightly during the COVID-19 pandemic. Great strides have been made to control and treat malaria—but not enough. New tools, like the ones being developed at CTEGD, are needed to keep pushing malaria’s morbidity and mortality rates in the right direction.
“The hard part—what can’t be done easily with the tools we already have—is being done,” Kyle said. “We just need new tools, which is one of the things that our center is really a leader in.”
Lọla Fagbami, a postdoctoral research associate at UGA, has been awarded a Burroughs Wellcome Fund 2022 Postdoctoral Diversity Enrichment Program fellowship.
Fagbami, UGA’s first PDEP Fellow, conducts research on the human malaria parasite Plasmodium falciparum at the Center for Tropical and Emerging Global Diseases. She works with Vasant Muralidharan, associate professor of cellular biology in the Franklin College of Arts and Sciences, who nominated her for the award.
“Dr. Fagbami has excellent training in metabolomics, mass spectrometry and Plasmodium drug discovery. Her exceptional work as a graduate student has shown how human malaria-causing parasites use metabolic adaptation to induce antimalarial drug resistance. Dr. Fagbami is a fearless, highly intelligent, accomplished and outstanding scientist who will be a leader in our field,” Muralidharan wrote in his nomination letter.
“Her research project addresses a major gap in the field that has enormous implications for malaria elimination and eradication efforts,” he added.
The PDEP award provides $60,000 over three years to support career-development activities for historically excluded minority postdoctoral fellows pursuing academic careers in biomedical or medical research, according to the Burroughs Wellcome Fund.
“This award is an investment in me as a scientist and leader and will help advance my career to the next level,” Fagbami said. “I am excited to join the extraordinary community of PDEP scholars and also connect with program alumni who have successfully made the transition to research independence.”
Fagbami earned a B.S. in chemistry at Emory University, an M.B.S. and an M.P.H. in health policy at Rutgers University, and a Ph.D. in chemical biology at Harvard University.
Malaria remains a major public health issue, infecting nearly 220 million people every year. The spread of drug-resistant strains of Plasmodium falciparum around the world threatens the progress made against this disease. Therefore, identifying druggable and essential pathways in P. falciparum parasites remains a major area of research. One poorly understood area of parasite biology is the formation of disulfide bonds, which is an essential requirement for the folding of numerous proteins. Specialized chaperones with thioredoxin (Trx) domains catalyze the redox functions necessary for breaking incorrect and forming correct disulfide bonds in proteins. Defining the substrates of these redox chaperones is difficult and immunoprecipitation based assays cannot distinguish between substrates and interacting partners. Further, the substrate or client interactions with the redox chaperones are usually transient in nature. Activity based crosslinkers that rely on the nucleophilic cysteines on Trx domains and the disulfide bond forming cysteines on clients provide an easily scalable method to trap and identify the substrates of Trx-domain containing chaperones. The cell permeable crosslinker divinyl sulfone (DVSF) is active only in the presence of nucleophilic cysteines in proteins and, therefore, traps Trx domains with their substrates, as they form mixed disulfide bonds during the course of their catalytic activity. This allows the identification of substrates that rely on Trx activity for their folding, as well as discovering small molecules that interfere with Trx domain activity. Graphic abstract: Identification of thioredoxin domain substrates via divinylsulfone crosslinking and immunoprecipitation-mass spectrometry.
David W Cobb, Grace S Woods, Vasant Muralidharan. Bio Protoc. 2022 Feb 20;12(4):e4322. doi: 10.21769/BioProtoc.4322.
Vasant Muralidharan, associate professor of cellular biology and a member of CTEGD, was recently interviewed for The Athens Frontline podcast episode The Next Pandemic. Listen as he discusses mRNA technology, the pandemic, and other emerging diseases.
