Biology subjects

Lyu, J.

Publications and source records attributed to Lyu, J..

At least 55 records · Page 3Linked to original sources

bioRxiv2023

A mitochondria-regulated p53-CCF circuit integrates genome integrity with inflammation

Genomic instability and inflammation are distinct hallmarks of aging, but the connection between them is poorly understood. Understanding their interrelationship will help unravel new mechanisms and therapeutic targets of aging and age-associated diseases. Here we report a novel mechanism directly linking genomic instability and inflammation in senescent cells through a mitochondria-regulated molecular circuit driven by p53 and cytoplasmic chromatin fragments (CCF). We show, through activation or inactivation of p53 by genetic and pharmacologic approaches, that p53 suppresses CCF accumulation and the downstream inflammatory senescence-associated secretory phenotype (SASP), without affecting cell cycle arrest. p53 activation suppressed CCF formation by promoting DNA repair, and this is reflected in maintenance of genomic integrity, particularly in subtelomeric regions, as shown by single cell genome resequencing. Activation of p53 in aged mice by pharmacological inhibition of MDM2 reversed signatures of aging, including age- and senescence-associated transcriptomic signatures of inflammation and age-associated accumulation of monocytes and macrophages in liver. Remarkably, mitochondria in senescent cells suppressed p53 activity by promoting CCF formation and thereby restricting ATM-dependent nuclear DNA damage signaling. These data provide evidence for a mitochondria-regulated p53 signaling circuit in senescent cells that controls DNA repair, genome integrity, and senescence- and age-associated inflammation. This pathway is immunomodulatory in mice and a potential target for healthy aging interventions by small molecules already shown to activate p53.

cell biology↗

bioRxiv2023

Protein migration and topography mapping reveals endogenous protein processing and homomers in Nicotiana benthamiana.

Processing by proteases irreversibly regulates the fate of plant proteins and hampers the production of recombinant protein in plants, yet only few processing events have been described in agroinfiltrated Nicotiana benthamiana, which has emerged as a favorite transient protein expression platform in plant science and molecular pharming. Here, we used in-gel digests and mass spectrometry to monitor the migration and topography of 5,040 plant proteins of agroinfiltrated N. benthamiana within a protein gel. By plotting the peptides over the gel slices, we generated peptographs that reveal where which part of each protein was detected within the protein gel. These data uncovered that 60% of the detected proteins have proteoforms that migrate at lower than predicted molecular weights, implicating extensive proteolytic processing. For instance, this analysis confirms the proteolytic removal and degradation of autoinhibitory prodomains of most but not all proteases, and revealed differential processing within pectinemethylesterase and lipase families. This analysis also uncovered intricate processing of glycosidases and uncovered that ectodomain shedding might be common for a diverse range of receptor-like kinases. Transient expression of double-tagged candidate proteins confirmed various processing events in vivo. This extensive proteomic dataset can be investigated further and demonstrates that most plant proteins are proteolytically processed and implicates an extensive proteolytic machinery shaping the proteome of agroinfiltrated N. benthamiana.

plant biology↗

bioRxiv2023

A peptide-centric local stability assay to unveil protein targets of diverse ligands

While tremendous progress has been made in chemical proteomics for identifying protein-ligand interactions, it remains challenging for proteome-wide identification of ligand-binding regions without modifying the ligands. Here, we discovered that "disruptive trypsinization" amplifies the readout of ligand-induced protein local stability shifts, and explored this notion in developing "peptide-centric local stability assay" (PELSA), a modification-free approach which achieves unprecedented sensitivity in proteome-wide target identification and binding-region determination. We demonstrate the versatility of PELSA by investigating the interactions across various biological contexts including drug-target interactions, metabolism, epitope mapping, metal proteomics, and post-translational modification recognition. A PELSA study of the oncometabolite R2HG revealed functional insights about its targets and pathogenic processes in both cancer and immune cells. Thus, beyond offering users unprecedented sensitivity for characterizing diverse target-ligand interactions, PELSA supports informative screening and hypothesis generation studies throughout life science.

biochemistry↗

bioRxiv2023

Mechanical force regulates ligand binding and function of PD-1

Immune checkpoint blockade targeting PD-1 shows great success in cancer therapy. However, the mechanism of how ligand binding initiates PD-1 signaling remains unclear. As prognosis markers of multiple cancers, soluble PD-L1 is found in patient sera and can bind PD-1, but fails to suppress T cell function. This and our previous observations that T cells exert endogenous forces on PD-1- PD-L2 bonds prompt the hypothesis that mechanical force might be critical to PD-1 triggering, which is missing in the soluble ligand case due to the lack of mechanical support afforded by surface-anchored ligand. Here we show that PD-1 function is eliminated or reduced when mechanical support on ligand is removed or dampened, respectively. Force spectroscopic analysis reveals that PD-1 forms catch bonds with both PD-Ligands <7 pN where force prolongs bond lifetime, but slip bonds >8 pN where force accelerates dissociation. Steered molecular dynamics finds PD-1-PD-L2 complex very sensitive to force due to the two molecules "side-to-side" binding via {beta} sheets. Pulling causes relative rotation and translation between the two molecules by stretching and aligning the complex along the force direction, yielding new atomic contacts not observed in the crystal structure. Compared to wild-type, PD-1 mutants targeting the force-induced new interactions maintain the same binding affinity but display lower rupture force, shorter bond lifetime, reduced tension, and most importantly, impaired capacity to suppress T cell activation. Our results uncover a mechanism for cells to probe the mechanical support of PD-1-PD-Ligand bonds using endogenous forces to regulate PD-1 triggering.

immunology↗

bioRxiv2023

Mechanotransduction governs CD40 function and underlies X-linked Hyper IgM syndrome

B cell maturation in germinal centers (GCs) depends on cognate interactions between the T and B cells. Upon interaction with CD40 ligand (CD40L) on T cells, CD40 delivers co-stimulatory signals alongside B cell antigen receptor (BCR) signaling to regulate affinity maturation and antibody class-switch during GC reaction. Mutations in CD40L disrupt interactions with CD40, which lead to abnormal antibody responses in immune deficiencies known as X-linked Hyper IgM syndrome (X-HIgM). Assuming that physical interactions between highly mobile T and B cells generate mechanical forces on CD40-CD40L bonds, we set out to study the B cell mechanobiology mediated by CD40-CD40L interaction. Using a suite of biophysical assays we find that CD40 forms catch bond with CD40L where the bond lasts longer at larger forces, B cells exert tension on CD40-CD40L bonds, and force enhances CD40 signaling and antibody class-switch. Significantly, X-HIgM CD40L mutations impair catch bond formation, suppress endogenous tension, and reduce force-enhanced CD40 signaling, leading to deficiencies in antibody class switch. Our findings highlight the critical role of mechanotransduction in CD40 function and provide insights into the molecular mechanisms underlying X-HIgM syndrome.

immunology↗

bioRxiv2023

Double and triple thermodynamic mutant cycles reveal the basis for specific MsbA-lipid interactions

Structural and functional studies of the ATP-binding cassette transporter MsbA have revealed two distinct lipopolysaccharide (LPS) binding sites: one located in the central cavity and the other at a membrane-facing, exterior site. Although these binding sites are known to be important for MsbA function, the thermodynamic basis for these specific MsbA-LPS interactions is not well understood. Here, we use native mass spectrometry to determine the thermodynamics of MsbA interacting with the LPS-precursor 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo)2-lipid A (KDL). The binding of KDL is solely driven by entropy, despite the transporter adopting an inward-facing conformation or trapped in an outward-facing conformation with adenosine 5-diphosphate and vanadate. An extension of the mutant cycle approach is employed to probe basic residues that interact with KDL. We find the molecular recognition of KDL is driven by a positive coupling entropy (as large as -100 kJ/mol at 298K) that outweighs unfavorable coupling enthalpy. These findings indicate that alterations in solvent reorganization and conformational entropy can contribute significantly to the free energy of protein-lipid association. The results presented herein showcase the advantage of native MS to obtain thermodynamic insight into protein-lipid interactions that would otherwise be intractable using traditional approaches, and this enabling technology will be instrumental in the life sciences and drug discovery.

biochemistry↗

bioRxiv2023

A marker chromosome in psychosis identifies glycine decarboxylase (GLDC) as a novel regulator of neuronal and synaptic function in the hippocampus

The biological significance of a small supernumerary marker chromosome that results in dosage alterations to chromosome 9p24.1, including triplication of the GLDC gene encoding glycine decarboxylase, in two patients with psychosis is unclear. In an allelic series of copy number variant mouse models, we identify that triplication of Gldc reduces extracellular glycine levels as determined by optical fluorescence resonance energy transfer (FRET) in dentate gyrus (DG) but not in CA1, suppresses long-term potentiation (LTP) in mPP-DG synapses but not in CA3-CA1 synapses, reduces the activity of biochemical pathways implicated in schizophrenia and mitochondrial bioenergetics, and displays deficits in prepulse inhibition, startle habituation, latent inhibition, working memory, sociability and social preference. Our results thus provide a link between a genomic copy number variation, biochemical, cellular and behavioral phenotypes, and further demonstrate that GLDC negatively regulates long-term synaptic plasticity at specific hippocampal synapses, possibly contributing to the development of neuropsychiatric disorders.

neuroscience↗

bioRxiv2023

Diazo-carboxyl click chemistry enables rapid and sensitive quantification of carboxylic acid metabolites

Carboxylic acids are central metabolites in bioenergetics, signal transduction and post-translation protein regulation. Unlike its genomic and transcriptomic counterparts, the quest for metabolomic profiling in trace amounts of biomedical samples is prohibitively challenging largely due to the lack of sensitive and robust quantification schemes for carboxylic acids. Based on diazo-carboxyl click chemistry, here we demonstrate DQmB-HA method as a rapid derivatization strategy for the sensitive analysis of hydrophilic, low-molecular-weight carboxylic acids. To the investigated metabolites, DQmB-HA derivatization method renders 5 to 2,000-fold higher response on mass spectrometry along with improved chromatographic separation on commercial UHPLC-MS machines. Using this method, we present the near-single-cell analysis of carboxylic acid metabolites in mouse egg cells before and after fertilization. Malate, fumarate and {beta}-hydroxybutyrate were found to decrease in mouse zygotes. We also showcase the kinetic profiling of TCA-cycle intermediates inside adherent cells cultured in one well of 96-well plates during drug treatment. FCCP and AZD3965 were shown to have overlapped but different effects on the isotope labeling of carboxylic acids. Finally, we apply DQmB-HA method to plasma or serum samples (down to 5 L) from mice and humans collected on pathological and physiological conditions. The measured changes of succinate, {beta}-hydroxybutyrate, and lactate in blood corroborate previous literatures in ischemia-reperfusion injury mouse model, acute fasting-refeeding mouse model, and human individuals diagnosed with mitochondrial dysfunction diseases, respectively. Overall, DQmB-HA method offers a sensitive, rapid and user-friendly quantification scheme for carboxylic acid metabolites, paving the road toward the ultimate goals of single-cell metabolomic analysis and bedside monitoring of biofluid samples.

systems biology↗

bioRxiv2023

LINE-associated cryptic splicing induces dsRNA-mediated interferon response and tumor immunity

RNA splicing plays a critical role in post-transcriptional gene regulation. Exponential expansion of intron length poses a challenge for accurate splicing. Little is known about how cells prevent inadvertent and often deleterious expression of intronic elements due to cryptic splicing. In this study, we identify hnRNPM as an essential RNA binding protein that suppresses cryptic splicing through binding to deep introns, preserving transcriptome integrity. Long interspersed nuclear elements (LINEs) harbor large amounts of pseudo splice sites in introns. hnRNPM preferentially binds at intronic LINEs and represses LINE-containing pseudo splice site usage for cryptic splicing. Remarkably, a subgroup of the cryptic exons can form long dsRNAs through base-pairing of inverted Alu transposable elements scattered in between LINEs and trigger interferon immune response, a well-known antiviral defense mechanism. Notably, these interferon-associated pathways are found to be upregulated in hnRNPM-deficient tumors, which also exhibit elevated immune cell infiltration. These findings unveil hnRNPM as a guardian of transcriptome integrity. Targeting hnRNPM in tumors may be used to trigger an inflammatory immune response thereby boosting cancer surveillance.

genomics↗

bioRxiv2023

Zika virus infection induces IL-1β-mediated inflammatory responses by macrophages in the brain of an adult mouse model

During the 2015/16 Zika virus (ZIKV) epidemic, ZIKV associated neurological diseases were reported in adults, including microcephaly, Guillain-Barre syndrome, myelitis, meningoencephalitis, and fatal encephalitis. However, the mechanisms underlying the neuropathogenesis of ZIKV infection are not yet fully understood. In this study, we used an adult ZIKV-infection mouse model (Ifnar1-/-) to investigate the mechanisms underlying neuroinflammation and neuropathogenesis. ZIKV infection induced the expression of proinflammatory cytokines, including IL-1{beta}, IL-6, IFN-{gamma}, and TNF-, in the brains of Ifnar1-/- mice. RNA-seq analysis of the infected mouse brain also revealed that genes involved in innate immune responses and cytokine-mediated signaling pathways were significantly upregulated at 6 days post infection. Furthermore, ZIKV infection induced macrophage infiltration and activation, and augmented IL-1{beta} expression, whereas microgliosis was not observed in the brain. Using human monocyte THP-1 cells, we confirmed that ZIKV infection promotes inflammatory cell death and increases IL-1{beta} secretion. In addition, the expression of complement component C3, which is associated with neurodegenerative diseases and known to be upregulated by proinflammatory cytokines, was induced by ZIKV infection through the IL-1{beta}-mediated pathway. An increase in C5a produced by complement activation in the brains of ZIKV-infected mice was also confirmed. Taken together, our results suggest that ZIKV infection of the brain in this animal model augments IL-1{beta} expression in infiltrating macrophages and elicits IL-1{beta}-mediated inflammation, which can lead to the destructive consequences of neuroinflammation. ImportanceZika virus (ZIKV) associated neurological impairments are an important global health problem. Our results suggest that ZIKV infection of the mouse brain can induce IL-1{beta}-mediated inflammation and complement activation, contributing to the development of neurological disorders. Thus, our findings reveal a mechanism by which ZIKV induces neuroinflammation in the mouse brain. Although we used adult type I IFN receptor IFNAR knockout (Ifnar1-/-) mice owing to the limited mouse model of ZIKV pathogenesis, our conclusion could contribute to understanding ZIKV associated neurological diseases to develop treatment strategies based on these findings for the patients with ZIKV infection.

microbiology↗

bioRxiv2022

From whole genome to probiotic candidates: a study of potential Lactobacillus strain selection for vaginitis treatment

Vaginitis is a syndrome characterized by not only the invasion of pathogens, but also the lack of Lactobacillus. Lactobacillus supplementation emerged as an innovative therapy for vaginitis which targeted the vaginal microbiota in recent years. However, limited live biotherapeutic products were explicitly marketed for this. The aim of our study is to characterize the potential Lactobacillus candidates for enhancing the treatment of vaginitis. Initially, 98 Lactobacillus isolates with whole genome were selected into the candidate pool. The genomic pathways involved in the biosynthesis of probiotic metabolites, adhering ability and acid/bile resistance, as well as sequences of antibiotic resistance genes, plasmids, transposons, viruses, and prophages were annotated. A scoring model was then established based on genomic analysis for ranking the performance of the strains, and the top-performing ones were applied to in vitro tests for sub-screening. As a result, two candidates were selected, and their probiotic abilities were verified with three reference strains. L. crispatus LG55-27 and L. gasseri TM13-16 presented outstanding ability to produce D-lactate and adhere to human vaginal epithelial cells. They also showed higher antimicrobial activity against Gardnerella vaginalis, Escherichia coli, Candida albicans, Staphylococcus aureus and Pseudomonas aeruginosa than control ones. Their acid/bile-resistant ability suggested the potential of oral supplementation. Strain LG55-27 and TM13-16 also display improved effects on pH changes, and the production of inflammation cytokines in BV model rats. This study provided a probiotic dosage recommendation for vaginitis treatment by demonstrating an effective probiotic screening pipeline based on genome sequences, in vitro tests and in vivo BV model experiment.

microbiology↗

bioRxiv2022

Genetic ablation of dentate hilar somatostatin-positive GABAergic interneurons is sufficient to induce cognitive impairment

Aging is often associated with a decline in cognitive function. A reduction in the number of somatostatin-positive (SOM+) interneurons in the dentate gyrus (DG) has been described in cognitively impaired but not in unimpaired aged rodents. However, it remains unclear whether the reduction in SOM+ interneurons in the DG hilus is causal for age-related cognitive dysfunction. We hypothesized that hilar SOM+ interneurons play an essential role in maintaining cognitive function and that a reduction in the number of hilar SOM+ interneurons might be sufficient to induce cognitive dysfunction. Hilar SOM+ interneurons were ablated by expressing a diphtheria toxin transgene specifically in these interneurons, which resulted in a reduction in the number of SOM+/GAD-67+ neurons and dendritic spine density in the DG. C-fos and Iba-1 immunostainings were increased in DG and CA3 but not in CA1. Behavioral testing revealed a reduced recognition index in the novel object recognition test, a reduction in the percentage of correct alternations in the Y maze tests, and increased latencies and path lengths in the learning and the reversal learning phase of the Morris water maze. Our results show that partial genetic ablation of SOM+ hilar interneurons is sufficient to increase activity in DG and CA3, as has been described to occur with aging and to induce an impairment of learning and memory functions. Thus, partial ablation of hilar SOM+ interneurons may be a significant contributing factor to age-related cognitive dysfunction. These mice may also be useful as a cellularly defined model of hippocampal aging.

neuroscience↗

bioRxiv2022

Chronic intermittent propofol attenuates surgery-induced neuroinflammation, apoptosis, and cognitive impairment in aged mice

Surgery may lead to long-lasting cognitive deficits that are referred to as perioperative neurocognitive disorder (NCD), particularly in elderly patients. Currently, no interventions are routinely employed in clinical practice to prevent perioperative NCD. Here we show that perioperative chronic intermittent administration of propofol to aged mice undergoing laparotomy under isoflurane anesthesia effectively blocks the surgery-induced increase in nitrosative stress, increased expression of proapoptotic proteins, microglial activation, and cognitive deficits. By contrast, in the absence of surgery and anesthesia, propofol had little effect on biochemical parameters and led to cognitive improvement only in a subset of behavioral paradigms. The actions of propofol were largely absent in mice lacking the GABAA receptor 5-subunit, indicating that they are mediated by 5-containing GABAA receptors. These results demonstrate that propofol - via 5-containing GABAA receptors that are redistributed to the cell surface membranes in a sustained manner - can attenuate surgery-induced neuroinflammation and postsurgical cognitive deficits.

neuroscience↗

bioRxiv2022

SCTC: inference of developmental potential from single-cell transcriptional complexity

Inference of single-cell developmental potential from scRNA-Seq data enables us to reconstruct the pseudo-temporal path of cell development, which is an important and challenging task for single-cell analysis. Single-cell transcriptional diversity (SCTD), measured by the number of expressed genes per cell, has been found to be negatively correlated with the development time, and thus can be considered as a hallmark of developmental potential. However, in some cases, the gene expression level of the cells in the early stages of development may be lower than that of the later stages, which may lead to incorrect estimation of differentiation states by gene diversity-based inference. Here we refer to the economic complexity theory and propose single-cell transcriptional complexity (SCTC) metrics as a measure of single-cell developmental potential, given the intrinsic similarities between biological and economic complex systems. We take into account not only the number of genes expressed by cells, but also the more sophisticated structure information of gene expression by treating the scRNA-seq count matrix as a bipartite network. We show that complexity metrics characterize the developmental potential more accurately than the diversity metrics. Especially, in the early stages of development, cells typically have lower gene expression level than that in the later stages, while their complexity in the early stages is significantly higher than that in the later stages. Based on the measurement of SCTC, we provide an unsupervised method for accurate, robust, and transferable inference of single-cell pseudotime. Our findings suggest that the complexity emerging from the interaction between cells and genes determines the developmental potential, which may bring new insights into the understanding of biological development from the perspective of the complexity theory.

bioinformatics↗

bioRxiv2022

Dissecting the cellular basis of age-related cognitive dysfunction: Chronic chemogenetic inhibition of somatostatin-positive interneurons in the dentate gyrus hilus induces memory impairments

The cellular basis of age-related impairments of hippocampal function is not fully understood. In order to evaluate the role of somatostatin-positive (Sst+) interneurons in the dentate gyrus hilus in this process, we chemogenetically inhibited Sst+ interneurons in the dentate gyrus (DG) hilus. Chronic chemogenetic inhibition (CCI) of these neurons resulted in increased c-Fos staining in the DG hilus, a decrease in the percentage of Gad67- and of Sst-expressing interneurons in the DG, and increased microglial activation in DG, CA3, and CA1. Total dendritic length and spine density were reduced in DG and CA1, suggesting reduced dendritic complexity. Behaviorally, the recognition index in an object recognition task and the percentage of spontaneous alternations in the Y maze were decreased, while in both initial and reversal learning in the Morris water maze the latencies to find the hidden platform were increased, suggesting cognitive dysfunction. Our findings establish a causal role for a reduced function of Sst+ interneurons in the DG hilus for cognitive decline and suggest that this reduced function may contribute to age-related impairments of learning and memory. Furthermore, our CCI mice may represent a cellularly defined model of hippocampal aging. SIGNIFICANCE STATEMENTNeuronal circuits and cellular processes underlying age-related cognitive dysfunction are not well understood. We observed that chronic chemogenetic inhibition of a defined cell type, somatostatin-positive interneurons in the dentate gyrus hilus, which have previously been found to be associated with cognitive dysfunction in aged rodents, is necessary and sufficient to elicit changes in expression of interneuronal markers, an increase in the activity of dentate gyrus granule cells, increased microglial activation across the entire hippocampus and an impairment of learning and memory-related tasks. Thus, inhibition of somatostatin-positive interneurons in the dentate gyrus hilus replicates changes that are also seen with normal aging, representing a novel cellularly defined animal model of hippocampal aging.

neuroscience↗

bioRxiv2022

Tumor microenvironments impair T cell receptor affinity and function

CD8+ T cells underpin effective anti-tumor immune responses in melanoma; however, their functions are attenuated due to various immunosuppressive factors in the tumor microenvironment (TME), resulting in disease progression. T cell function is elicited by the T cell receptor (TCR), which recognizes antigen peptide-major histocompatibility complex (pMHC) expressed on tumor cells via direct physical contact, i.e., two-dimensional (2D) interaction. TCR-pMHC 2D affinity plays a central role in antigen recognition and discrimination, and is sensitive to both the conditions of the T cell and the microenvironment in which it resides. Herein, we demonstrate that CD8+ T cells residing in TME have lower 2D TCR-pMHC bimolecular affinity and TCR-pMHC-CD8 trimolecular avidity, pull fewer TCR-pMHC bonds by endogenous forces, flux lower level of intracellular calcium in response to antigen stimulation, exhibit impaired in vivo activation, and show diminished anti-tumor effector function. These detrimental effects are localized in the tumor and tumor draining lymph node (TdLN), and affect both antigen-inexperienced and antigen-experienced CD8+ T cells irrespective of their TCR specificities. These findings implicate impaired antigen recognition as a mechanism of T cell dysfunction in the TME.

immunology↗

bioRxiv2022

Structural basis for lipid and copper regulation of the ABC transporter MsbA

A critical step in Lipopolysaccharide (LPS) biogenesis involves flipping lipooligosaccharide, an LPS precursor, from the cytoplasmic to the periplasmic leaflet of the inner membrane, an operation carried out by the ATP-binding cassette transporter MsbA. Although MsbA has been extensively studied, the selectivity of MsbA-lipid interactions remains poorly understood. Here we use native mass spectrometry (MS) to characterize MsbA-lipid interactions and guide structural studies. We show the transporter co-purifies with copper(II) and metal binding modulates protein-lipid interactions. A 2.15 [A] resolution structure of an N-terminal region of MsbA in complex with copper(II) is presented, revealing a structure reminiscent of the GHK peptide, a high-affinity copper(II) chelator. Our results demonstrate conformation-dependent lipid binding affinities, particularly for the LPS-precursor, 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo)2-lipid A (KLA). We report a 3.6 [A]-resolution structure of MsbA trapped in an open, outward-facing conformation with adenosine 5-diphosphate and vanadate, revealing an unprecedented KLA binding site, wherein the lipid forms extensive interactions with the transporter. Additional studies provide evidence that the exterior KLA binding site is conserved and a positive allosteric modulator of ATPase activity, serving as a feedforward activation mechanism to couple transporter activity with LPS biosynthesis.

biochemistry↗

bioRxiv2022

Large library docking for novel SARS-CoV-2 main protease non-covalent and covalent inhibitors

Antiviral therapeutics to treat SARS-CoV-2 are much desired for the on-going pandemic. A well-precedented viral enzyme is the main protease (MPro), which is now targeted by an approved drug and by several investigational drugs. With the inevitable liabilities of these new drugs, and facing viral resistance, there remains a call for new chemical scaffolds against MPro. We virtually docked 1.2 billion non-covalent and a new library of 6.5 million electrophilic molecules against the enzyme structure. From these, 29 non-covalent and 11 covalent inhibitors were identified in 37 series, the most potent having an IC50 of 29 M and 20 M, respectively. Several series were optimized, resulting in inhibitors active in the low micromolar range. Subsequent crystallography confirmed the docking predicted binding modes and may template further optimization. Together, these compounds reveal new chemotypes to aid in further discovery of MPro inhibitors for SARS-CoV-2 and other future coronaviruses.

biophysics↗