Abstract Title
Mutational Analysis of Human Norovirus VP2 Elucidates Critical Molecular Interactions for Virus Assembly
Presenter
Sue Crawford, Baylor College of Medicine
Co-Author(s)
Janam Dave [1,2], Sue E. Crawford [2] , Khalil Ettayebi [2], B.V.V Prasad [1,3], Mary K. Estes [1,2,4]1-Chemical, Physical and Structural Biology Graduate Program 2-Department of Molecular Virology & Microbiology, 3-Department of Biochemistry and Molecular Pharmacology, 4-Department of Medicine, Baylor College of Medicine, Houston, TX
Abstract Category
Structure & Pathogenesis
Abstract
Human noroviruses (HuNoVs) are the leading global cause of viral gastroenteritis, resulting in 680 million cases annually, with > 80% of infections caused by the GII genogroup. HuNoVs are non-enveloped, with an outer capsid composed of VP1 which encloses a minor structural protein, VP2, and a (+) ssRNA genome covalently attached to a translation initiator protein, VPg. An atomic resolution structure of VP1 is known through X-ray crystallography and cryo-EM analyses of HuNoV VLPs, but the structure of VP2 and VPg inside the capsid are unresolved. Therefore, the biochemical nature of VP1, VP2 and VPg’s interactions and how the virus particle assembles are fundamental knowledge gaps.
Herein, we show VP2 is the molecular bridge required for complete assembly. We used virus like particles (VLPs), deletion constructs, and mutational analyses to determine the interaction site on VP2 for VP1 of the globally prevalent and pandemic-causing GII.4 Sydney HuNoV. We identified amino acid (AA) residues 40-43 on VP2 are required for interaction with VP1 and mutation of VP2 AA 40-43 abrogates VP2 encapsidation. Computational analysis predicts VP2 has a highly conserved N-terminal alpha helical domain and an intrinsically disordered C-terminal domain. Co-variation evolutionary trace analysis indicates the VP1 shell domain has co-evolved with the N-terminal domain of VP2, especially at interacting residues. Next, we identified VP2, not VP1, uniquely binds VPg and recruits VPg using its C-terminal domain. These findings uncover the fundamentals of HuNoV assembly which is mediated by VP2, highlighting VP2 as a putative antiviral target.
Herein, we show VP2 is the molecular bridge required for complete assembly. We used virus like particles (VLPs), deletion constructs, and mutational analyses to determine the interaction site on VP2 for VP1 of the globally prevalent and pandemic-causing GII.4 Sydney HuNoV. We identified amino acid (AA) residues 40-43 on VP2 are required for interaction with VP1 and mutation of VP2 AA 40-43 abrogates VP2 encapsidation. Computational analysis predicts VP2 has a highly conserved N-terminal alpha helical domain and an intrinsically disordered C-terminal domain. Co-variation evolutionary trace analysis indicates the VP1 shell domain has co-evolved with the N-terminal domain of VP2, especially at interacting residues. Next, we identified VP2, not VP1, uniquely binds VPg and recruits VPg using its C-terminal domain. These findings uncover the fundamentals of HuNoV assembly which is mediated by VP2, highlighting VP2 as a putative antiviral target.