Residue-level resolution of alphavirus envelope protein interactions in pH-dependent fusion

Abstract

Alphavirus envelope proteins, organized as trimers of E2-E1 heterodimers on the surface of the pathogenic alphavirus, mediate the low pH-triggered fusion of viral and endosomal membranes in human cells. The lack of specific treatment for alphaviral infections motivates our exploration of potential antiviral approaches by inhibiting one or more fusion steps in the common endocytic viral entry pathway. In this work, we performed constant pH molecular dynamics based on an atomic model of the alphavirus envelope with icosahedral symmetry. We have identified pH-sensitive residues that cause the largest shifts in thermodynamic driving forces under neutral and acidic pH conditions for various fusion steps. A series of conserved interdomain His residues is identified to be responsible for the pH-dependent conformational changes in the fusion process, and ligand binding sites in their vicinity are anticipated to be potential drug targets aimed at inhibiting viral infections. 3). The lack of a vaccine or specific treatment prompts investigations of the fundamental mechanisms of the alphaviral lifecycle to facilitate the development of effective antiviral therapies (4). Alphaviruses have been reported to enter the cell through receptor-mediated endocytosis. Here, alphaviruses are ferried toward the perinuclear space of the host cell inside vesicles towed by molecular motors and delivered to specific locations for productive replication (5-11). Even when direct entry into the cytoplasm is possible (11-15), the endocytic entry pathway facilitates the transportation of viruses across the crowded cytoplasmic space and delays detection by the immune system without leaving empty capsid or envelope as obvious evidence of the viral infection exposed outside the host cell (10, 11). Before the delivery of its viral genome into the cytoplasm of a host cell, the alphavirus must undergo a critical step of low pH-triggered membrane fusion, which is a common mechanism in the endocytic viral entry pathway among many different viruses. Understanding the mechanism of the low pH-triggered alphaviral membrane fusion is essential for the development of therapies against alphavirus as well as other viruses using similar endocytic entry mechanisms.Recent studies of the lifecycle of alphavirus reveal that a precursor, p62, is first synthesized as a chaperon forming a heterodimer with E1, which is essential for viral budding (16); p62 protects the E1 protein in the low-pH environment of the secretory pathway before being cleaved by cellular furin to produce mature E2-E1 and a smaller fragment, E3 (17-21). After the virus buds from the cytoplasmic membrane, E3 is released from the virus particle under neutral pH conditions outside the host cell (13,(22)(23)(24).On the surface of a mature alphavirus, 80 (E2-E1) 3 viral spikes, organized in T = 4 icosahedral symmetry on the viral lipid membrane, enclose the viral capsid and genome (25-43). On internalization of the mature virus in the endosome of the host cell in a new round of infection cycle, the ...

CHARMM‐GUI 10 years for biomolecular modeling and simulation

“…For example, CHARMM-GUI currently supports a system building up to 3 million atoms, but as larger spatial scales, longer time scales, and higher levels of realism are becoming possible and necessary, we plan to dynamically adjust this limit in the future. In addition, more modules will be available in Input Generator for advanced simulations techniques, such as QM and QM/MM calculations, [121][122][123][124] NMR structure calculation and refinement, [125][126][127][128] multisite kdynamics, [129][130][131] constant pH simulations, [132][133][134][135][136][137][138][139][140][141][142][143] grid-based energy correction map, [144,145] and various replica-exchange enhanced sampling methods. [146] In terms of system building, our effort will be placed toward making Glycolipid Modeler and LPS Modeler available in Membrane Builder and other modules, enabling crowding and cellular-scale system modeling, and extending CHARMM-GUI for nanoscale material sciences.…”

Unraveling HIV protease flaps dynamics by Constant pH Molecular Dynamics simulations

“…The currently available CpHMD methods can be fundamentally categorized into two main approaches: continuous protonation states (Goh et al, 2014;Lee et al, 2004;May et al, 2014;Shen, 2011 andZeng et al, 2015) and discrete protonation states (Bürgi et al, 2002;Itoh et al, 2011;McGee et al, 2014;Mongan et al, 2004 andSwails et al, 2014).…”

“…The most widely employed approach regarding these models is the Constant pH Molecular Dynamics (CpHMD) (Bürgi et al, 2002 andMongan et al, 2004), in which the solution pH should be specified as an external variable. One advantage of these CpHMD methods is that they allow the investigation of pH-dependent properties of proteins, and the correlation between protonation and conformational states (Dissanayake et al, 2015;Shi et al, 2012 andZeng et al, 2015).…”

Insights into Adenovirus Uncoating from Interactions with Integrins and Mediators of Host Immunity

“…Precisely what factor or factors trigger adenovirus conformational changes within the endosome is unknown. Constant-pH molecular dynamics simulations for alphaviruses and bacteriophage HK97 have identified specific residues in these viruses that respond to changes in pH and lead to a shifting of the stability between two conformational states [17,18]. In the case of adenovirus, it is possible that the low pH of the endosome triggers a conformational change, or alternatively perhaps ions or enzymes in high concentration in the endosome are responsible.…”