scholarly journals Small-Molecule Inhibition of Viral Fusion Glycoproteins

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Han-Yuan Liu ◽  
Priscilla L. Yang
Keyword(s):  
Membrane Fusion ◽  
Small Molecule ◽  
Viral Entry ◽  
Small Molecule Inhibitors ◽  
Fusion Reaction ◽  
Covalent Bond ◽  
Annual Review ◽  
Publication Date ◽  
Viral Fusion ◽  
Viral Glycoprotein

Viral fusion glycoproteins catalyze membrane fusion during viral entry. Unlike most enzymes, however, they lack a conventional active site in which formation or scission of a specific covalent bond is catalyzed. Instead, they drive the membrane fusion reaction by cojoining highly regulated changes in conformation to membrane deformation. Despite the challenges in applying inhibitor design approaches to these proteins, recent advances in knowledge of the structures and mechanisms of viral fusogens have enabled the development of small-molecule inhibitors of both class I and class II viral fusion proteins. Here, we review well-validated inhibitors, including their discovery, targets, and mechanism(s) of action, while highlighting mechanistic similarities and differences. Together, these examples make a compelling case for small-molecule inhibitors as tools for probing the mechanisms of viral glycoprotein-mediated fusion and for viral glycoproteins as druggable targets. Expected final online publication date for the Annual Review of Virology, Volume 8 is September 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

scholarly journals New Biophysical Approaches Reveal the Dynamics and Mechanics of Type I Viral Fusion Machinery and Their Interplay with Membranes

Viruses ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 413 ◽  
Author(s):  
Mark A. Benhaim ◽  
Kelly K. Lee
Keyword(s):  
Membrane Fusion ◽  
Conformational Changes ◽  
Viral Entry ◽  
Fusion Proteins ◽  
Structural Changes ◽  
Fusion Reaction ◽  
Type I ◽  
Viral Fusion ◽  
Technological Advances ◽  
Viral Fusion Proteins

Protein-mediated membrane fusion is a highly regulated biological process essential for cellular and organismal functions and infection by enveloped viruses. During viral entry the membrane fusion reaction is catalyzed by specialized protein machinery on the viral surface. These viral fusion proteins undergo a series of dramatic structural changes during membrane fusion where they engage, remodel, and ultimately fuse with the host membrane. The structural and dynamic nature of these conformational changes and their impact on the membranes have long-eluded characterization. Recent advances in structural and biophysical methodologies have enabled researchers to directly observe viral fusion proteins as they carry out their functions during membrane fusion. Here we review the structure and function of type I viral fusion proteins and mechanisms of protein-mediated membrane fusion. We highlight how recent technological advances and new biophysical approaches are providing unprecedented new insight into the membrane fusion reaction.

scholarly journals How small-molecule inhibitors of dengue-virus infection interfere with viral membrane fusion

2018 ◽  
Author(s):  
Luke H. Chao ◽  
Jaebong Jang ◽  
Adam Johnson ◽  
Anthony Nguyen ◽  
Nathanael S. Gray ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Membrane Fusion ◽  
Small Molecule ◽  
Single Particle ◽  
Small Molecule Inhibitors ◽  
Viral Fusion ◽  
Viral Attachment ◽  
Target Membrane ◽  
Viral Membrane Fusion ◽  
Rate Limiting

AbstractDengue virus (DV) is a compact, icoshedrally symmetric, enveloped particle, covered by 90 dimers of envelope protein (E), which mediates viral attachment and membrane fusion. Fusion requires a dimer-to-trimer transition and membrane engagement of hydrophobic “fusion loops”. We previously characterized the steps in membrane fusion for the related West Nile virus (WNV), using recombinant, WNV virus-like particles (VLPs) for single-particle experiments. Trimerization and membrane engagement are rate-limiting; fusion requires at least two adjacent trimers; availability of competent monomers within the contact zone between virus and target membrane creates a trimerization bottleneck. We have extended that work to dengue VLPs, from all four DV serotypes, finding an essentially similar mechanism. Small-molecule inhibitors of DV infection that target E block its fusion-inducing conformation change. We show that ∼15 bound molecules per particle (∼8.5 % occupancy) completely prevent fusion, in accord with the proposed mechanism and the likely inhibitor binding site on E.Impact statementSingle-particle studies of dengue-virus membrane fusion and the effect of small-molecule inhibitors of infection clarify the viral fusion mechanism.

scholarly journals Small Molecule Inhibitors of Influenza Virus Entry

2021 ◽  
Vol 14 (6) ◽  
pp. 587
Author(s):  
Zhaoyu Chen ◽  
Qinghua Cui ◽  
Michael Caffrey ◽  
Lijun Rong ◽  
Ruikun Du
Keyword(s):  
Influenza Virus ◽  
Membrane Fusion ◽  
Small Molecule ◽  
Drug Target ◽  
Small Molecule Inhibitors ◽  
Virus Entry ◽  
Critical Role ◽  
Structural Basis ◽  
Future Directions ◽  
The Past

Hemagglutinin (HA) plays a critical role during influenza virus receptor binding and subsequent membrane fusion process, thus HA has become a promising drug target. For the past several decades, we and other researchers have discovered a series of HA inhibitors mainly targeting its fusion machinery. In this review, we summarize the advances in HA-targeted development of small molecule inhibitors. Moreover, we discuss the structural basis and mode of action of these inhibitors, and speculate upon future directions toward more potent inhibitors of membrane fusion and potential anti-influenza drugs.

scholarly journals pH-Induced Activation of Arenavirus Membrane Fusion Is Antagonized by Small-Molecule Inhibitors

2008 ◽  
Vol 82 (21) ◽  
pp. 10932-10939 ◽  
Author(s):  
Joanne York ◽  
Dongcheng Dai ◽  
Sean M. Amberg ◽  
Jack H. Nunberg
Keyword(s):  
Membrane Fusion ◽  
Small Molecule ◽  
Viral Entry ◽  
De Novo ◽  
Lassa Fever ◽  
Molecular Target ◽  
Active Inhibitor ◽  
Hemorrhagic Fevers ◽  
Lassa Fever Virus ◽  
Small Molecule Compound

ABSTRACT The arenavirus envelope glycoprotein (GPC) mediates viral entry through pH-induced membrane fusion in the endosome. This crucial process in the viral life cycle can be specifically inhibited in the New World arenaviruses by the small-molecule compound ST-294. Here, we show that ST-294 interferes with GPC-mediated membrane fusion by targeting the interaction of the G2 fusion subunit with the stable signal peptide (SSP). We demonstrate that amino acid substitutions at lysine-33 of the Junín virus SSP confer resistance to ST-294 and engender de novo sensitivity to ST-161, a chemically distinct inhibitor of the Old World Lassa fever virus. These compounds, as well as a broadly active inhibitor, ST-193, likely share a molecular target at the SSP-G2 interface. We also show that both ST-294 and ST-193 inhibit pH-induced dissociation of the G1 receptor-binding subunit from GPC, a process concomitant with fusion activation. Interestingly, the inhibitory activity of these molecules can in some cases be overcome by further lowering the pH used for activation. Our results suggest that these small molecules act to stabilize the prefusion GPC complex against acidic pH. The pH-sensitive interaction between SSP and G2 in GPC represents a robust molecular target for the development of antiviral compounds for the treatment of arenavirus hemorrhagic fevers.

scholarly journals Small Molecule Inhibitors in the Treatment of Rheumatoid Arthritis and Beyond: Latest Updates and Potential Strategy for Fighting COVID-19

Cells ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 1876
Author(s):  
Magdalena Massalska ◽  
Wlodzimierz Maslinski ◽  
Marzena Ciechomska
Keyword(s):  
Rheumatoid Arthritis ◽  
Small Molecule ◽  
Viral Entry ◽  
Small Molecule Inhibitors ◽  
Large Family ◽  
Downstream Signaling ◽  
Wide Range ◽  
Synthetic Dmards ◽  
Potential Strategy ◽  
Inflammatory Molecules

The development of biological disease-modifying antirheumatic drugs (bDMARDs) and target synthetic DMARDs (tsDMARDs), also known as small molecule inhibitors, represent a breakthrough in rheumatoid arthritis (RA) treatment. The tsDMARDs are a large family of small molecules targeting mostly the several types of kinases, which are essential in downstream signaling of pro-inflammatory molecules. This review highlights current challenges associated with the treatment of RA using small molecule inhibitors targeting intracellular JAKs/MAPKs/NF-κB/SYK-BTK signaling pathways. Indeed, we have provided the latest update on development of small molecule inhibitors, their clinical efficacy and safety as a strategy for RA treatment. On the other hand, we have highlighted the risk and adverse effects of tsDMARDs administration including, among others, infections and thromboembolism. Therefore, performance of blood tests or viral infection screening should be recommended before the tsDMARDs administration. Interestingly, recent events of SARS-CoV-2 outbreak have demonstrated the potential use of small molecule inhibitors not only in RA treatment, but also in fighting COVID-19 via blocking the viral entry, preventing of hyperimmune activation and reducing cytokine storm. Thus, small molecule inhibitors, targeting wide range of pro-inflammatory singling pathways, may find wider implications not only for the management of RA but also in the controlling of COVID-19.

scholarly journals A Small-molecule Inhibitor Directed against the Chemokine Receptor CXCR4 Prevents its Use as an HIV-1 Coreceptor

1997 ◽  
Vol 186 (8) ◽  
pp. 1395-1400 ◽  
Author(s):  
Benjamin J. Doranz ◽  
Kathie Grovit-Ferbas ◽  
Matthew P. Sharron ◽  
Si-Hua Mao ◽  
Matthew Bidwell Goetz ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Small Molecule ◽  
Chemokine Receptor ◽  
Viral Entry ◽  
Small Molecule Inhibitor ◽  
Chemokine Receptor Cxcr4 ◽  
Mechanism Of Inhibition ◽  
Molecule Inhibitor ◽  
Immunodeficiency Virus ◽  
Hiv 1

The chemokine receptor CXCR4 is the major coreceptor used for cellular entry by T cell– tropic human immunodeficiency virus (HIV)-1 strains, whereas CCR5 is used by macrophage (M)-tropic strains. Here we show that a small-molecule inhibitor, ALX40-4C, inhibits HIV-1 envelope (Env)-mediated membrane fusion and viral entry directly at the level of coreceptor use. ALX40-4C inhibited HIV-1 use of the coreceptor CXCR4 by T- and dual-tropic HIV-1 strains, whereas use of CCR5 by M- and dual-tropic strains was not inhibited. Dual-tropic viruses capable of using both CXCR4 and CCR5 were inhibited by ALX40-4C only when cells expressed CXCR4 alone. ALX40-4C blocked stromal-derived factor (SDF)-1α–mediated activation of CXCR4 and binding of the monoclonal antibody 12G5 to cells expressing CXCR4. Overlap of the ALX40-4C binding site with that of 12G5 and SDF implicates direct blocking of Env interactions, rather than downregulation of receptor, as the mechanism of inhibition. Thus, ALX40-4C represents a small-molecule inhibitor of HIV-1 infection that acts directly against a chemokine receptor at the level of Env-mediated membrane fusion.

scholarly journals An Epitope of the Semliki Forest Virus Fusion Protein Exposed during Virus-Membrane Fusion

1999 ◽  
Vol 73 (12) ◽  
pp. 10029-10039 ◽  
Author(s):  
Anna Ahn ◽  
Matthew R. Klimjack ◽  
Prodyot K. Chatterjee ◽  
Margaret Kielian
Keyword(s):  
Membrane Fusion ◽  
Binding Site ◽  
Semliki Forest Virus ◽  
Fusion Reaction ◽  
Ph Dependence ◽  
Fusion Peptide ◽  
Endocytic Pathway ◽  
Spike Protein ◽  
Viral Fusion ◽  
Wide Range

ABSTRACT Semliki Forest virus (SFV) is an enveloped alphavirus that infects cells via a membrane fusion reaction triggered by acidic pH in the endocytic pathway. Fusion is mediated by the spike protein E1 subunit, an integral membrane protein that contains the viral fusion peptide and forms a stable homotrimer during fusion. We have characterized four monoclonal antibodies (MAbs) specific for the acid conformation of E1. These MAbs did not inhibit fusion, suggesting that they bind to an E1 region different from the fusion peptide. Competition analyses demonstrated that all four MAbs bound to spatially related sites on acid-treated virions or isolated spike proteins. To map the binding site, we selected for virus mutants resistant to one of the MAbs, E1a-1. One virus isolate, SFV 4-2, showed reduced binding of three acid-specific MAbs including E1a-1, while its binding of one acid-specific MAb as well as non-acid-specific MAbs to E1 and E2 was unchanged. The SFV 4-2 mutant was fully infectious, formed the E1 homotrimer, and had the wild-type pH dependence of infection. Sequence analysis demonstrated that the relevant mutation in SFV 4-2 was a change of E1 glycine 157 to arginine (G157R). Decreased binding of MAb E1a-1 was observed under a wide range of assay conditions, strongly suggesting that the E1 G157R mutation directly affects the MAb binding site. These data thus localize an E1 region that is normally hidden in the neutral pH structure and becomes exposed as part of the reorganization of the spike protein to its fusion-active conformation.

scholarly journals A Virion-Based Assay for Glycoprotein Thermostability Reveals Key Determinants of Filovirus Entry and Its Inhibition

2020 ◽  
Vol 94 (18) ◽  
Author(s):  
Robert H. Bortz ◽  
Anthony C. Wong ◽  
Michael G. Grodus ◽  
Hannah S. Recht ◽  
Marc C. Pulanco ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Small Molecule ◽  
Viral Entry ◽  
Ebola Virus ◽  
Elevated Temperatures ◽  
Vaccine Development ◽  
Cysteine Proteases ◽  
Enzyme Linked Immunosorbent Assay ◽  
Temperature Dependent ◽  
Acid Ph

ABSTRACT Ebola virus (EBOV) entry into cells is mediated by its spike glycoprotein (GP). Following attachment and internalization, virions traffic to late endosomes where GP is cleaved by host cysteine proteases. Cleaved GP then binds its cellular receptor, Niemann-Pick C1. In response to an unknown cellular trigger, GP undergoes conformational rearrangements that drive fusion of viral and endosomal membranes. The temperature-dependent stability (thermostability) of the prefusion conformers of class I viral fusion glycoproteins, including those of filovirus GPs, has provided insights into their propensity to undergo fusion-related rearrangements. However, previously described assays have relied on soluble glycoprotein ectodomains. Here, we developed a simple enzyme-linked immunosorbent assay (ELISA)-based assay that uses the temperature-dependent loss of conformational epitopes to measure thermostability of GP embedded in viral membranes. The base and glycan cap subdomains of all filovirus GPs tested suffered a concerted loss of prefusion conformation at elevated temperatures but did so at different temperature ranges, indicating virus-specific differences in thermostability. Despite these differences, all of these GPs displayed reduced thermostability upon cleavage to GP conformers (GPCL). Surprisingly, acid pH enhanced, rather than decreased, GP thermostability, suggesting it could enhance viral survival in hostile endo/lysosomal compartments. Finally, we confirmed and extended previous findings that some small-molecule inhibitors of filovirus entry destabilize EBOV GP and uncovered evidence that the most potent inhibitors act through multiple mechanisms. We establish the epitope-loss ELISA as a useful tool for studies of filovirus entry, engineering of GP variants with enhanced stability for use in vaccine development, and discovery of new stability-modulating antivirals. IMPORTANCE The development of Ebola virus countermeasures is challenged by our limited understanding of cell entry, especially at the step of membrane fusion. The surface-exposed viral protein, GP, mediates membrane fusion and undergoes major structural rearrangements during this process. The stability of GP at elevated temperatures (thermostability) can provide insights into its capacity to undergo these rearrangements. Here, we describe a new assay that uses GP-specific antibodies to measure GP thermostability under a variety of conditions relevant to viral entry. We show that proteolytic cleavage and acid pH have significant effects on GP thermostability that shed light on their respective roles in viral entry. We also show that the assay can be used to study how small-molecule entry inhibitors affect GP stability. This work provides a simple and readily accessible assay to engineer stabilized GP variants for antiviral vaccines and to discover and improve drugs that act by modulating GP stability.

scholarly journals The Polar Region of the HIV-1 Envelope Protein Determines Viral Fusion and Infectivity by Stabilizing the gp120-gp41 Association

2019 ◽  
Vol 93 (7) ◽  
Author(s):  
Wuxun Lu ◽  
Shuliang Chen ◽  
Jingyou Yu ◽  
Ryan Behrens ◽  
Joshua Wiggins ◽  
...  
Keyword(s):  
Membrane Fusion ◽  
Cell Fusion ◽  
Viral Entry ◽  
Polar Region ◽  
Targeted Mutagenesis ◽  
Target Cells ◽  
Viral Fusion ◽  
Underlying Mechanisms ◽  
Hiv 1

ABSTRACT HIV-1 enters cells through binding between viral envelope glycoprotein (Env) and cellular receptors to initiate virus and cell fusion. HIV-1 Env precursor (gp160) is cleaved into two units noncovalently bound to form a trimer on virions, including a surface unit (gp120) and a transmembrane unit (gp41) responsible for virus binding and membrane fusion, respectively. The polar region (PR) at the N terminus of gp41 comprises 17 residues, including 7 polar amino acids. Previous studies suggested that the PR contributes to HIV-1 membrane fusion and infectivity; however, the precise role of the PR in Env-mediated viral entry and the underlying mechanisms remain unknown. Here, we show that the PR is critical for HIV-1 fusion and infectivity by stabilizing Env trimers. Through analyzing the PR sequences of 57,645 HIV-1 isolates, we performed targeted mutagenesis and functional studies of three highly conserved polar residues in the PR (S532P, T534A, and T536A) which have not been characterized previously. We found that single or combined mutations of these three residues abolished or significantly decreased HIV-1 infectivity without affecting viral production. These PR mutations abolished or significantly reduced HIV-1 fusion with target cells and also Env-mediated cell-cell fusion. Three PR mutations containing S532P substantially reduced gp120 and gp41 association, Env trimer stability, and increased gp120 shedding. Furthermore, S532A mutation significantly reduced HIV-1 infectivity and fusogenicity but not Env expression and cleavage. Our findings suggest that the PR of gp41, particularly the key residue S532, is structurally essential for maintaining HIV-1 Env trimer, viral fusogenicity, and infectivity. IMPORTANCE Although extensive studies of the transmembrane unit (gp41) of HIV-1 Env have led to a fusion inhibitor clinically used to block viral entry, the functions of different domains of gp41 in HIV-1 fusion and infectivity are not fully elucidated. The polar region (PR) of gp41 has been proposed to participate in HIV-1 membrane fusion in biochemical analyses, but its role in viral entry and infectivity remain unclear. In our effort to characterize three nucleotide mutations of an HIV-1 RNA element that partially overlaps the PR coding sequence, we identified a novel function of the PR that determines viral fusion and infectivity. We further demonstrated the structural and functional impact of six PR mutations on HIV-1 Env stability, viral fusion, and infectivity. Our findings reveal the previously unappreciated function of the PR and the underlying mechanisms, highlighting the important role of the PR in regulating HIV-1 fusion and infectivity.

scholarly journals The Membrane-Proximal Domain of Vesicular Stomatitis Virus G Protein Functions as a Membrane Fusion Potentiator and Can Induce Hemifusion

2002 ◽  
Vol 76 (23) ◽  
pp. 12300-12311 ◽  
Author(s):  
E. Jeetendra ◽  
Clinton S. Robison ◽  
Lorraine M. Albritton ◽  
Michael A. Whitt
Keyword(s):  
Membrane Fusion ◽  
G Protein ◽  
Vesicular Stomatitis Virus ◽  
Fusion Proteins ◽  
Fold Increase ◽  
Wild Type Virus ◽  
Viral Fusion ◽  
Fusion Activity ◽  
Viral Glycoprotein ◽  
Vesicular Stomatitis

ABSTRACT Recently we showed that the membrane-proximal stem region of the vesicular stomatitis virus (VSV) G protein ectodomain (G stem [GS]), together with the transmembrane and cytoplasmic domains, was sufficient to mediate efficient VSV budding (C. S. Robison and M. A. Whitt, J. Virol. 74:2239-2246, 2000). Here, we show that GS can also potentiate the membrane fusion activity of heterologous viral fusion proteins when GS is coexpressed with those proteins. For some fusion proteins, there was as much as a 40-fold increase in syncytium formation when GS was coexpressed compared to that seen when the fusion protein was expressed alone. Fusion potentiation by GS was not protein specific, since it occurred with both pH-dependent as well as pH-independent fusion proteins. Using a recombinant vesicular stomatitis virus encoding GS that contained an N-terminal hemagglutinin (HA) tag (GSHA virus), we found that the GSHA virus bound to cells as well as the wild-type virus did at pH 7.0; however, the GSHA virus was noninfectious. Analysis of cells expressing GSHA in a three-color membrane fusion assay revealed that GSHA could induce lipid mixing but not cytoplasmic mixing, indicating that GS can induce hemifusion. Treatment of GSHA virus-bound cells with the membrane-destabilizing drug chlorpromazine rescued the hemifusion block and allowed entry and subsequent replication of GSHA virus, demonstrating that GS-mediated hemifusion was a functional intermediate in the membrane fusion pathway. Using a series of truncation mutants, we also determined that only 14 residues of GS, together with the VSV G transmembrane and cytoplasmic tail, were sufficient for fusion potentiation. To our knowledge, this is the first report which shows that a small domain of one viral glycoprotein can promote the fusion activity of other, unrelated viral glycoproteins.

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