Tag: molecular biology

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  • Researchers identify a new approach to controlling bacterial infections

    Researchers identify a new approach to controlling bacterial infections

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    Researchers at the Icahn School of Medicine at Mount Sinai have identified a new approach to controlling bacterial infections. The findings were described in the February 6 online issue of Nature Structural & Molecular Biology [DOI # 10.1038/s41594-024-01220-x].

    The team found a way to turn on a vital bacterial defense mechanism to fight and manage bacterial infections. The defense system, called cyclic oligonucleotide-based antiphage signaling system (CBASS), is a natural mechanism used by certain bacteria to protect themselves from viral attacks. Bacteria self-destruct as a means to prevent the spread of virus to other bacterial cells in the population.

    We wanted to see how the bacterial self-killing CBASS system is activated and whether it can be leveraged to limit bacterial infections. This is a fresh approach to tackling bacterial infections, a significant concern in hospitals and other settings. It’s essential to find new tools for fighting antibiotic resistance. In the war against superbugs, we need to constantly innovate and expand our toolkit to stay ahead of evolving drug resistance.”


    Aneel Aggarwal, PhD, Co-Senior Author, Professor of Pharmacological Sciences at Icahn Mount Sinai

    According to a 2019 report by the Centers for Disease Control and Prevention, more than 2.8 million antimicrobial-resistant infections occur in the United States each year, with over 35,000 people dying as a result.

    As part of the experiments, the researchers studied how “Cap5,” or CBASS-associated protein 5, is activated for DNA degradation and how it could be used to control bacterial infections through a combination of structural analysis and various biophysical, biochemical, and cellular assays. Cap5 is a key protein that becomes activated by cyclic nucleotides (small signaling molecules) to destroy the bacterial cell’s own DNA.

    “In our study, we started by identifying which of the many cyclic nucleotides could activate the effector Cap5 of the CBASS system,” says co-senior author Olga Rechkoblit, PhD, Assistant Professor of Pharmacological Sciences at Icahn Mount Sinai. “Once we figured that out, we looked closely at the structure of Cap5 when it’s bound to these small signaling molecules. Then, with expert help from Daniela Sciaky, PhD, a researcher at Icahn Mount Sinai, we showed that by adding these special molecules to the bacteria’s environment, these molecules could potentially be used to eliminate the bacteria.”

    The researchers found that determining the structure of Cap5 with cyclic nucleotides posed a technical challenge, requiring expert help from Dale F. Kreitler, PhD, AMX Beamline Scientist at Brookhaven National Laboratory. It was achieved by using micro-focused synchrotron X-ray radiation at the same facility. Micro-focused synchrotron X-ray radiation is a type of X-ray radiation that is not only produced using a specific type of particle accelerator (synchrotron) but is also carefully concentrated or focused on a tiny area for more detailed imaging or analysis.

    Next, the researchers will explore how their discoveries apply to other types of bacteria and assess whether their method can be used to manage infections caused by various harmful bacteria.

    The paper is titled “Activation of CBASS-Cap5 endonuclease immune effector by cyclic nucleotides.”

    Other authors who contributed to this work are Angeliki Buku, PhD, and Jithesh Kottur, PhD, both with Icahn Mount Sinai.

    The work was funded by National Institutes of Health grants R35-GM131780, P41GM111244, KP1605010, P30 GM124165, S10OD021527, GM103310, and by the Simons Foundation grant SF349247.

    Source:

    Journal reference:

    Rechkoblit, O., et al. (2024). Activation of CBASS Cap5 endonuclease immune effector by cyclic nucleotides. Nature Structural & Molecular Biology. doi.org/10.1038/s41594-024-01220-x.

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  • Unraveling the architecture of poxvirus cores

    Unraveling the architecture of poxvirus cores

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    A recent re-emergence and outbreak of Mpox brought poxviruses back as a public health threat, underlining an important knowledge gap at their core. Now, a team of researchers from the Institute of Science and Technology Austria (ISTA) lifted the mysteries of poxviral core architecture by combining various cryo-electron microscopy techniques with molecular modeling. The findings, published in Nature Structural & Molecular Biology, could facilitate future research on therapeutics targeting the poxvirus core.

    Variola virus, the most notorious poxvirus and one of the deadliest viruses to have afflicted humans, wreaked havoc by causing smallpox until it was eradicated in 1980. The eradication succeeded thanks to an extensive vaccination campaign using another poxvirus, the aptly named Vaccinia virus. The 2022-2023 re-emergence and outbreak of Mpox virus reminded us once more that viruses find ways to return to the forefront as public health threats. Importantly, this has highlighted the fundamental questions about poxviruses that have remained unanswered to this day.

    One such fundamental question lies, quite literally, at the core of the matter: “We know that for poxviruses to be infective, their viral core must be properly formed. But what is this poxviral core made of, and how do its individual components come together and function?” asks ISTA Assistant Professor Florian Schur, the corresponding author of the study.

    Schur and his team now put their finger on the missing link: a protein called A10. Interestingly, A10 is common to all clinically relevant poxviruses. In addition, the researchers found that A10 acts as one of the main building blocks of the poxviral core. This knowledge could be instrumental for future research on therapeutics targeting the poxviral core.

    “The most advanced cryo-EM techniques available today”

    The viral core is one of the factors common to all infectious poxvirus forms.

    Previous experiments in virology, biochemistry, and genetics suggested several core protein candidates for poxviruses, but there were no experimentally-derived structures available.”


    Julia Datler, ISTA PhD student, one of the co-first authors of the study

    Thus, the team started by computationally predicting models of the main core protein candidates, using the now-famous AI-based molecular modeling tool AlphaFold. In parallel, Datler was setting the project’s biochemical and structural foundations by drawing on her background in virology and the Schur group’s main expertise: cryogenic electron microscopy, or cryo-EM for short. “We integrated many of the most advanced cryo-EM techniques available today with AlphaFold molecular modeling. This gave us, for the first time, a detailed overall view of the poxviral core–the ‘safe’ or ‘bioreactor’ inside the virus that encloses the viral genome and releases it in infected cells,” says Schur. “It was a bit of a gamble, but we eventually managed to find the right mix of techniques to examine this complex question,” says postdoc Jesse Hansen, the study’s co-first author whose expertise in various structural biology techniques and image processing methods was pivotal for the project.

    A global 3D view of the poxvirus

    The ISTA researchers examined “live” Vaccinia virus mature virions and purified poxviral cores under every possible angle–quite literally. “We combined the ‘classic’ single-particle cryo-EM, cryo-electron tomography, subtomogram averaging, and AlphaFold analysis to gain an overall view of the poxviral core,” says Datler. With cryo-electron tomography, researchers can reconstitute 3D volumes of a biological sample as large as an entire virus by acquiring images while gradually tilting the sample. “It’s like doing a CT scan of the virus,” says Hansen. “Cryo-electron tomography, our lab’s ‘specialty,’ allowed us to gain nanometer-level resolutions of the whole virus, its core, and interior,” says Schur. In addition, the researchers could fit the AlphaFold models into the observed shapes like a puzzle and identify molecules that make up the poxviral core. Among these, the core protein candidate A10 stood out as one of the major components. “We found that A10 defines key structural elements of the core of poxviruses,” says Datler. Schur adds, “These findings are a great resource to interpret bits of structural and virological data generated over the last decades.”

    A rugged path to uncovering poxviral cores

    The path to these findings was all but straightforward. “We needed to find our own way from the start,” says Datler. Leveraging her expertise in biochemistry, virology, and structural biology, Datler isolated, propagated, and purified samples of Vaccinia virus and established the protocols to purify the complete viral core, all while optimizing these samples for structural studies. “Structurally, it was extremely hard to study these virus cores. But luckily, our perseverance and optimism paid off,” says Hansen.

    The ISTA researchers are convinced that their findings could provide a knowledge platform for future therapeutics that seek to target poxviral cores. “For example, one could think of drugs that prevent the core from assembling – or even disassembling and releasing the viral DNA during infection. Ultimately, fundamental virus research, as done here, allows us to be better prepared against possible future viral outbreaks,” concludes Schur.

    Source:

    Journal reference:

    Datler, J., et al. (2024). Multi-modal cryo-EM reveals trimers of protein A10 to form the palisade layer in poxvirus cores. Nature Structural & Molecular Biology. doi.org/10.1038/s41594-023-01201-6.

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  • NIH awards $1.9 million to investigate how complex communities of microbes impact human health

    NIH awards $1.9 million to investigate how complex communities of microbes impact human health

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    The bacteria and microorganisms that live within the human body -; the microbiota -; can impact human health, disease risk and even how the body absorbs medications, but the details of these processes are unclear. To help understand how complex communities of microbes impact human health, the National Institutes of Health’s National Institute of General Medical Science has awarded a 5-year, $1.9 million Maximizing Investigator’s Research Award to Jordan Bisanz, assistant professor of biochemistry and molecular biology in the Penn State Eberly College of Science.

    It is clear that the diversity of microbes in the human body is important, but why that diversity is important gets much murkier. We plan to use a combination of computational and experimental biology to better understand the different species that make up a healthy human microbiome as well as how they work together to impact human health.” 


    Jordan Bisanz, assistant professor of biochemistry and molecular biology, Penn State Eberly College of Science

    A single person’s microbiome might contain several hundred or thousand different species, for example living within the digestive tract or on the skin. Many microbiome studies focus on counting the number of different species present -; the diversity -; as well as identifying individual species and their abundance. Bisanz plans to use machine learning and other computational methods to analyze these kinds of publicly available data to better understand what defines a healthy microbial community.

    “We plan to look at information from hundreds of people to identify the most common strains or combinations of strains in a healthy microbiome,” he said. “Then, we can essentially design a healthy microbiome and use that as a model system to understand how the community functions. This is a very different approach from how microbiology tends to be done.”

    After identifying the key species in a typical healthy microbiome, Bisanz and his lab will experimentally build a laboratory model of a healthy microbiome, pulling from the several hundred microbial strains they maintain in the lab. Then, they can assess how that microbiome might respond in the context of a certain disease. They could also identify and build laboratory models microbiomes that reflect those of individuals with particular diseases.

    “It’s kind of like a cookbook of microbial communities, and we can decide what is practical for us to use to explore different questions,” he said. “Ultimately, we hope to better identify the specific microbes and metabolic processes that are important for host-microbe interactions and how they impact disease risk.”

    This work could help guide the development of targeted therapies, Bisanz said, that use specific microbes to treat bacterial infections or to improve gut health, rather than a more generic set of microbes, like those in probiotics and fecal transplant therapy.

    Bisanz will also explore how the microbiome might impact a person’s response to medications. Most drugs taken orally pass through microbes before they are absorbed in the body, he said, so microbes -; like host genetics -; can impact how effectively and efficiently drugs are absorbed. 

    “One of the questions from a personalized medicine standpoint is how to select the right dosage of a drug,” he said. “You can imagine in a futuristic world, you might go to the doctor for a treatment and they could do a quick test. And maybe they would identify a genetic marker that indicates you absorb the drug more quickly than others and also identify that they have a microbe that is known to metabolize the drug, and that helps them set the dosage.”

    Bisanz and colleagues will specifically investigate the role of the microbiome in the uptake of orally administered antimalarial drugs, which are notorious for their side effects that range from dizziness and vomiting to lucid dreams.

    “There is a huge amount of unexplained variation in the amount and severity of these side-effects, and a lot of people stop taking the drugs because of those side effects,” Bisanz said. “Antimalarials are also a subject of concern because there is increasing resistance to the most common drugs in the parasites that cause malaria. If microbes can impact how much of a drug gets into the bloodstream, they may also influence the side effects, and it’s reasonable that they may also be playing a role in the development of resistance.”

    Together, this grant will provide Bisanz the opportunity to improve understanding of how microbial communities shape human health and physiology. 

    “The microbiome is very responsive, and it can be modified quickly and easily using dietary manipulations, like probiotics,” he said. “It is a relatively young field, but it has promising and immediate translational applications.”

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  • Inside the secretive cannabis lab that makes futuristic weed

    Inside the secretive cannabis lab that makes futuristic weed

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    Cannabis is changing. Today’s marijuana is more potent than ever and there remain many questions about its benefits and risks. We still can’t meaningfully regulate driving while high. And growing cannabis – legally and not – has a staggering environmental footprint.

    In The weed of the future, the final episode of our special three-part podcast series on the science of cannabis, Christie Taylor steps back to examine how cannabis fits into our society, both now and in the future, and how scientists may be able to breed cannabis with less of a tiny molecule that is responsible for the…

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