SARS-CoV-2 protein caught severing critical immunity pathway

Powerful X-rays from SLAC’s synchrotron reveal that our immune system’s primary wiring seems to be no match for a brutal SARS-CoV-2 protein.

BY DAVID KRAUSE

Over the past two years, scientists have studied the SARS-CoV-2 virus in great detail, laying the foundation for developing COVID-19 vaccines and antiviral treatments. Now, for the first time, scientists at the Department of Energy’s SLAC National Accelerator Laboratory have seen one of the virus’s most critical interactions, which could help researchers develop more precise treatments.

The team caught the moment when a virus protein, called Mpro, cuts a protective protein, known as NEMO, in an infected person. Without NEMO, an immune system is slower to respond to increasing viral loads or new infections. Seeing how Mpro attacks NEMO at the molecular level could inspire new therapeutic approaches.

To see how Mpro cuts NEMO, researchers funneled powerful X-rays from SLAC’s Stanford Synchrotron Radiation Lightsource (SSRL) onto crystallized samples of the protein complex. The X-rays struck the protein samples, revealing what Mpro looks like when it dismantles NEMO’s primary function of helping our immune system communicate.

“We saw that the virus protein cuts through NEMO as easily as sharp scissors through thin paper,” said co-senior author Soichi Wakatsuki, professor at SLAC and Stanford. “Imagine the bad things that happen when good proteins in our bodies start getting cut into pieces.”

The images from SSRL show the exact location of NEMO’s cut and provide the first structure of SARS-CoV-2 Mpro bound to a human protein.

“If you can block the sites where Mpro binds to NEMO, you can stop this cut from happening over and over,” SSRL lead scientist and co-author Irimpan Mathews said. “Stopping Mpro could slow down how fast the virus takes over a body. Solving the crystal structure revealed Mpro’s binding sites and was one of the first steps to stopping the protein.”

The research team from SLAC, DOE’s Oak Ridge National Laboratory, and other institutions published their results today in Nature Communications.

Read more on the SLAC website

A promising treatment for ovarian cancer

Scientists are looking to harness the immune system to fight cancer

Over 20,000 women across the U.S. and Canada are diagnosed with ovarian cancer annually. The symptoms of this disease are often overlooked until it has spread, making it difficult to detect and treat with conventional methods like radiation and chemotherapy.

Dr. Cory Books, Associate Professor in the Department of Chemistry and Biochemistry at California State University, Fresno, is looking to harness the immune system to fight cancer. He is interested in a particular protein, called mucin, that is found throughout the body and is involved with the production of mucus. This protein is altered in cancer cells, which makes it a unique target for researchers.

“The cell stops adding sugars to the protein, so instead of having this mucus layer, now it has a solid protein layer, and cancer uses that to help spread itself through the body,” Brooks said.

This alteration helps ovarian cancer grow and spread, but it also leaves a signal that can help clinicians locate the cancer and kill it.

“What that means now is that there’s sort of this unique signature that we can target with antibodies to develop a new treatment for cancer,” Brooks said.

Researchers have been interested in this protein since the late 1980s but have never before been able to visualize how antibodies interact with the molecule.

With the help of the CMCF beamline at the Canadian Light Source (CLS) located at the University of Saskatchewan, Brooks and his team were able to see how antibodies bind to the protein for the first time.

Read more on the CLS website

Image: Brandy White, lead author on the study and graduate student with the Department of Chemistry and Biochemistry at California State University, Fresno.

How to get chloride ions into the cell

For the first time, a molecular movie has captured in detail the process of an anion transported across the cell membrane by a light-fueled protein pump. Publishing in Science, the researchers utilized the unique synergy of a Free Electron Laser (SwissFEL) and synchrotron light source (SLS) offered by PSI to unravel the mystery of how light energy initiates the pumping process − and how nature made sure there is no anion leakage back outside.

Many bacteria and unicellular algae have light-driven pumps in their cell membranes: proteins that change shape when exposed to photons such that they can transport charged atoms in or out of the cell. Thanks to these pumps, their unicellular owners can adjust to the environment’s pH value or salinity.

One such bacteria is Nonlabens marinus, first discovered in 2012 in the Pacific Ocean. Among others, it possesses a rhodopsin protein in its cell membrane which transports chloride anions from outside the cell to its inside. Just like in the human eye, a retinal molecule bound to the protein isomerizes when exposed to light. This isomerization starts the pumping process. Researchers now gained detailed insight into how the chloride pump in Nonlabens marinus works.

The study was led by Przemyslaw Nogly, once a postdoc at PSI and now an Ambizione Fellow and Group Leader at ETH Zürich, in close collaboration with the ALVRA team at SwissFEL and the MX team at the SLS. It is one of the first studies to fully combine experimental capabilities at these large-scale research facilities, bridging the gap in time resolution to record a full molecular movie of a protein at work. Slower dynamics in the millisecond-range were investigated via time-resolved serial crystallography at SLS while faster, up to picosecond, events were captured at SwissFEL – then both sets of data were put together.

Read more the PSI website

Image: Photoactive chloride pumping through the cell membrane captured by time-resolved serial crystallography: Chloride ions (green spheres) are transported across the cell membrane by the NmHR chloride pump (pink).

Credit: Guillaume Gotthard, Sandra Mous

Light sources have demonstrated huge adaptability during the pandemic

Johanna Hakanpää is the beamline scientist for P11, one of the macromolecular crystallography beamlines at PETRAIII at DESY in Hamburg. Originally from Finland, she studied chemistry and then did her masters and PhD work in protein crystallography. Johanna was drawn to the field because she wanted to understand how life really works. Supporting health related research is important to her and Johanna is especially inspired by her son who is a patient of celiac disease. Together they hope that one day, with the help of science, he will be able to eat normally without having to think about what is contained in his food. Johanna started her light source journey as a user and was really impressed by the staff scientists who supported her during her experiments. This led her to apply for a beamline scientist position and she successfully made the transition, learning the technical aspects of the beamlines on the job.

In her #LightSourceSelfie, Johanna highlights the adaptability of light sources during the pandemic as a key strength. Being part of a team that was able to keep the lights on for users via remote experiments is a reflection of the commitment that Johanna and her colleagues have when it comes to facilitating science. Thousands of staff at light sources all around the world have shown the same commitment, ensuring scientific advances can continue. This is particularly true for vital research on the SARS-CoV-2 virus itself. Learn more about this research here: https://lightsources.org/lightsource-research-and-sars-cov-2/

APS helps Pfizer create Covid-19 antiviral treatment

Pharmaceutical company Pfizer has announced the results of clinical trials of its new oral antiviral treatment against COVID-19. The new drug candidate, Paxlovid, proved to be effective against the SARS-CoV-2 virus, which causes COVID-19, according to results released by Pfizer on Nov. 5.

Scientists at Pfizer created Paxlovid with the help of the ultrabright X-rays of the Advanced Photon Source (APS), a U.S. Department of Energy (DOE) Office of Science user facility at DOE’s Argonne National Laboratory.

“Today’s news is a real game-changer in the global efforts to halt the devastation of this pandemic,” said Albert Bourla, chairman and chief executive officer of Pfizer, in a company press release. ​“These data suggest that our oral antiviral candidate, if approved or authorized by regulatory authorities, has the potential to save patients’ lives, reduce the severity of COVID-19 infections and eliminate up to nine out of 10 hospitalizations.”

DOE invests in user facilities such as the APS for the benefit of the nation’s scientific community, and supports biological research as part of its energy mission. This research has been critical in the fight against COVID-19. The DOE national laboratories formed the National Virtual Biotechnology Laboratory (NVBL) consortium in 2020 to combat COVID-19 using capabilities developed for their DOE mission, and that consortium helps support research into antiviral treatments such as Paxlovid.

Work to determine the structure of the antiviral candidate was done at the Industrial Macromolecular Crystallography Association Collaborative Access Team (IMCA-CAT) beamline at the APS, operated by the Hauptman-Woodward Medical Research Institute (HWI) on behalf of a collaboration of pharmaceutical companies, of which Pfizer is a member.

As a member of IMCA-CAT, Pfizer routinely conducts drug development experiments at the APS, and the process of narrowing down and zeroing in on this drug candidate was performed over many months, according to Lisa Keefe, executive director of IMCA-CAT and vice president for advancing therapeutics and principal scientist at Hauptman-Woodward Medical Research Institute. IMCA-CAT, she said, delivers quality results in a timely manner, much faster than the home laboratories of the companies themselves can do.

Read more on the APS website

Image: The IMCA-CAT beamline at the Advanced Photon Source, where work was done to determine the structure of Pfizer’s new COVID-19 antiviral treatment candidate.

Credit: Lisa Keefe, IMCA-CAT/Hauptman-Woodward Medical Research Institute

#LightSourceSelfies – Light Source scientists are innovators

Kathryn Janzen is an Associate Scientist and User Experience Coordinator at the Canadian Light Source. During her #LightSourceSelfie, Kathryn reflects on the light source community saying “The contacts between light sources are really important and everyone is very interested in sharing ideas. We’re also really interested in innovating and finding new ways to use the light source and finding new applications for old techniques.”

Understanding how a key antibody targets cancer cells

Immunotherapy can be used as a precise intervention in cancer treatments. Jean-Philippe Julien is a Canada Research Chair in Structural Immunology, a Senior Scientist in the Molecular Medicine Program at The Hospital for Sick Children (SickKids), and an Associate Professor in the Departments of Biochemistry and Immunology at the University of Toronto. Along with colleagues from the U.S., Spain and Canada, he used the Canadian Light Source at the University of Saskatchewan to study how a candidate antibody therapeutic interacts with a surface receptor on cancer cells, which provides important molecular insights for designing improved cancer therapies. He mentioned how the synchrotron is “incredibly important for researchers like myself” and how “we cannot do the research that we do without it.” The team used the CMCF beamline at the CLS and their findings were published in the Journal of Biological Chemistry.Immunotherapy can be used as a precise intervention in cancer treatments. Jean-Philippe Julien is a Canada Research Chair in Structural Immunology, a Senior Scientist in the Molecular Medicine Program at The Hospital for Sick Children (SickKids), and an Associate Professor in the Departments of Biochemistry and Immunology at the University of Toronto. Along with colleagues from the U.S., Spain and Canada, he used the Canadian Light Source at the University of Saskatchewan to study how a candidate antibody therapeutic interacts with a surface receptor on cancer cells, which provides important molecular insights for designing improved cancer therapies. He mentioned how the synchrotron is “incredibly important for researchers like myself” and how “we cannot do the research that we do without it.” The team used the CMCF beamline at the CLS and their findings were published in the Journal of Biological Chemistry.

Learn more on the CLS website

Image: Jean-Philippe Julien

Credit: Canadian Light Source

Giving rice new weapons to fight rice blast disease

Understanding how a fungal pathogen interacts with rice cells could help us engineer new defences 

Rice is one of the world’s most important agricultural crops, with 741.5 million tonnes produced in 2014. A large proportion of the global population relies on rice as a staple food, particularly in Asia and Africa. However, harvests are threatened by rice blast disease, caused by the fungus Magnaporthe oryzae, which destroys enough rice to feed around 200 million people every year. Rice and the rice blast fungus are involved in a co-evolutionary arms race, fighting for the upper hand. As the fungus relies on effector proteins to help it infect and reproduce within rice plants, rice has evolved immune receptors that allow it to detect and prevent the spread of the fungus. However, the rice blast fungus has evolved stealthy effector proteins that remain undetected by the rice immune system but can still promote disease. In work recently published in the Journal of Biological Chemistry, an international team of scientists has investigated how one stealthy effector protein might maintain its disease-promoting activity but evade immune detection. This research has an ultimate aim of engineering a receptor that would allow rice plants to better defend themselves. 

A pain in the paddy field

We’re familiar with images of the rice paddies of Asia, but this impressive sight represents an irresistible target for the rice blast fungus, Magnaporthe oryzae. Unable to run away from pests and pathogens, plants have evolved immune systems to detect and defend against attack. However, huge swathes planted with the same variety creates an evolutionary pressure for pests and pathogens; a feast is at hand if they can evade those defences. 

One way that pathogens try and gain an advantage is through the use of effector proteins. These proteins can suppress the plant’s immune system and manipulate the plant’s own systems to help the pathogen infect and replicate. However, the mechanisms they employ to do so are not fully understood.  

In collaboration with scientists from Japan and Thailand, researchers at the UK’s John Innes Centre and The Sainsbury Laboratory have been investigating the interaction between rice plants and the rice blast fungus, with the ultimate goal of engineering new genetic resources that will help rice fight this damaging disease.

Read more on the Diamond website

Image: Rice fields in Asia

A novel approach offers hope for an HCV vaccine

An HCV vaccine is needed, but hard to develop. A structural mimic may be the key to enhancing our immune response

Globally, more than 70 million people were struggling with a chronic hepatitis C virus (HCV) infection in 2015. Although effective drugs are available to treat chronic infections, only 13% of cases received curative treatment. The fact that only 20% have been diagnosed is of even greater concern. Although a minority of newly-infected individuals (10–40%) manage to overcome the disease, most develop a chronic infection. Most acute cases of HCV are asymptomatic, leading to undetected virus transmission. Left untreated chronic HCV can lead to serious liver damage and an increased risk of liver cancer. As curative therapies alone cannot eliminate the virus, a vaccine is required. However, because HCV is very diverse and evolves rapidly to evade the immune system, developing an effective vaccine is challenging. In work recently published in npj Vaccines, scientists from the MRC-University of Glasgow Centre for Virus Research, the University of St. Andrews and Imperial College London describe an alternative strategy that uses a structural mimic to encourage the immune system to make antibodies that can recognise multiple strains of the virus i.e. broadly-neutralising antibodies (bNAbs) against HCV. 

A moving target

With its high genetic diversity and an envelope of ever-changing glycoproteins, HCV is challenging for the human immune system to detect and counteract. The minority of cases in which the virus is successfully cleared from the body show a broad, strong T-cell response and neutralising antibodies during the early phase of infection. Individuals who have previously cleared an HCV infection have an 80% chance of successfully fighting off reinfection, indicating that a protective immune response has been induced and that vaccination is a realistic goal. However, with seven distinct genotypes and more than 60 subtypes, the genetic variation makes it challenging to produce a vaccine that would protect against all infections. 

Read more on the Diamond website

Image: I03 beamline at Diamond

Credit: Diamond Light Source

Understanding what makes COVID-19 more infectious than SARS

Australian and International researchers continue to have rapid access to the macromolecular and microfocus beamlines at the Australian Synchrotron to solve protein structures in the fight against COVID-19.

“Since coming out of a hard lockdown, we are now accepting proposals for other research,” said Principal Scientist Dr Alan Riboldi-Tunnicliffe.

“Because scientists can access the beamline remotely, they do not have to worry about changes to borders and travel restrictions.”

There have been a number of COVID-19 publications, which included structural information about key proteins in the virus, from the beamlines.

Instrument scientist Dr Eleanor Campbell reports that an international team of researchers led by the University of Bristol (UK) have identified a possible cause of SARS-CoV-2’s increased infectivity compared to SARS-CoV (the virus which emerged in China in 2003) , which could provide a target for developing COVID-19 therapies.

Australian collaborators included researchers from the Institute of Molecular Bioscience at the University of Queensland, who sent the samples to the Australian Synchrotron.

Read more on the Australian Synchrotron website

Scientists discover potential method to starve the bacteria that cause Tuberculosis

By deepening our understanding of how Tuberculosis bacteria feed themselves, University of Guelph researchers have identified a potential target for drug treatment. The team used the Canadian Light Source (CLS) at the University of Saskatchewan to image the bacteria in fine detail.

The infectious disease Tuberculosis (TB) is one of the leading causes of death worldwide. While rates of TB in Canada have remained relatively static since the 1980s, the disease disproportionately affects Indigenous populations. With TB-causing bacteria becoming increasingly resistant to antibiotics, researchers and drug makers are eager to find new, more effective treatments.

Researchers have known for some time that the bacteria that causes TB (Mycobacterium tuberculosis) uses our body’s cholesterol – a steroid – as a food source. Other relatives of the bacteria that do not cause disease share its ability to break down steroids. In this study, the University of Guelph team identified the structure of an enzyme (acyl CoA dehydrogenase) involved in steroid degradation in another member of the same bacteria family, called Thermomonospora curvata.

Read more on the CLS Website

Image: This rendering shows the shape of a tunnel (orange) where the substrate binds. Any drugs targeting this enzyme would need to fit to this pocket.

“foot-2-foot” interaction sheds light on bacterial conjugation

Bacteria possess mechanisms to establish communication between cells. This is especially important in bacterial conjugation, a process that allows bacteria to share genetic material. This is often used by bacteria to transfer antibiotic resistance genes and other virulence factors to neighbor cells, increasing the antibiotic resistance spread.

Now, a research team of ALBA scientists report the structural mechanism by which two proteins, Rap and Rco, act together to regulate conjugation. Rco is a repressor of conjugation, whereas Rap binds Rco and prevents Rco-mediated conjugation repression, thus resulting in an activation of the conjugation mechanism. The main results of the study show that Rap contains a binding pocket were a short peptide can bind, producing structural changes in Rap that forces its tetramerization, releasing Rco for blocking conjugation. Tetramerization occurs through an interaction that scientists named “foot-2-foot”, which differs significantly from the model proposed for other proteins of the Rap family.

Read more on the ALBA website

Image: RappLS20 tetramerization, side view of the peptide-bound tetramer. The red arrows indicate the loops connecting helices H4 and H5. (C) Zoom of the area around the N-terminus of helix H4, showing the insertion of this helix into the opposite monomer. The homotetramerization caused by the foot-2-foot interactions of the NTDs of RappLS20 provides an explanation for the activation of the RcopLS20 partner. In the absence of the peptide, the NTDs are positioned such that they allow the interaction with RcopLS20. However, upon binding the signaling peptide, the NTDs shift outwards, facilitating the formation of the homotetramer, leading to a change of the interaction surface of the NTDs that is no longer available for interactions with RcopLS20

Effective new target for breast cancer treatment

An international study led by scientists at the University of Sussex has provided strong evidence for an effective new target for breast cancer treatment. The five-year study, called “The structure-function relationship of oncogenic LMTK3” published in Science Advances, involved researchers from seven institutions across three countries including Diamond. 

The study suggests that LMTK3 inhibitors could be effectively used for the treatment of breast cancer, and potentially other types of cancer. The structure of oncogenic LMTK3 (Lemur Tyrosine Kinase 3 ) determines its role and functions allowing drug inhibition as a new therapeutic strategy.

It is hoped the research will allow the further development and optimisation of LMTK3 inhibitors as a new type of orally-administered anticancer drug for patients and have potential value not only for breast cancer patients but also for lung, stomach, thyroid and bladder cancer patients.

Read more on the Diamond Light Source website

Image: Crystal structure of LMTK3
Credit: University of Sussex