title: An Optimized Hepatitis C Virus E2 Glycoprotein Core Adopts a Functional Homodimer That Efficiently Blocks Virus Entry.
abstract: The hepatitis C virus (HCV) envelope glycoprotein E2 is the major target of broadly neutralizing antibodiesin vivoand is the focus of efforts in the rational design of a universal B cell vaccine against HCV. The E2 glycoprotein exhibits a high degree of amino acid variability which localizes to three discrete regions: hypervariable region 1 (HVR1), hypervariable region 2 (HVR2), and the intergenotypic variable region (igVR). All three variable regions contribute to immune evasion and/or isolate-specific structural variations, both important considerations for vaccine design. A high-resolution structural definition of the intact HCV envelope glycoprotein complex containing E1 and E2 remains to be elucidated, while crystallographic structures of a recombinant E2 ectodomain failed to resolve HVR1, HVR2, and a major neutralization determinant adjacent to HVR1. To obtain further information on E2, we characterized the role of all three variable regions in E2 ectodomain folding and function in the context of a recombinant ectodomain fragment (rE2). We report that removal of the variable regions accelerates binding to the major host cell receptor CD81 and that simultaneous deletion of HVR2 and the igVR is required to maintain wild-type CD81-binding characteristics. The removal of the variable regions also rescued the ability of rE2 to form a functional homodimer. We propose that the rE2 core provides novel insights into the role of the variable motifs in the higher-order assembly of the E2 ectodomain and may have implications for E1E2 structure on the virion surface.IMPORTANCEHepatitis C virus (HCV) infection affects ∼2% of the population globally, and no vaccine is available. HCV is a highly variable virus, and understanding the presentation of key antigenic sites at the virion surface is important for the design of a universal vaccine. This study investigates the role of three surface-exposed variable regions in E2 glycoprotein folding and function in the context of a recombinant soluble ectodomain. Our data demonstrate the variable motifs modulate binding of the E2 ectodomain to the major host cell receptor CD81 and have an impact on the formation of an E2 homodimer with high-affinity binding to CD81.
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Australia.|Department of Microbiology and Immunology, The University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia.|Cellular Protein Chemistry, Utrecht University, Utrecht, The Netherlands.
authors.forename: Kathleen
authors.initials: K
authors.lastname: McCaffrey
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Australia.
authors.forename: Irene
authors.initials: I
authors.lastname: Boo
authors.affiliation: Research and Development, CSL Limited, Parkville, Victoria, Australia.
authors.forename: Catherine M
authors.initials: CM
authors.lastname: Owczarek
authors.affiliation: Research and Development, CSL Limited, Parkville, Victoria, Australia.
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Hardy
authors.affiliation: Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria, Australia.
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Perugini
authors.affiliation: Research and Development, CSL Limited, Parkville, Victoria, Australia.
authors.forename: Louis
authors.initials: L
authors.lastname: Fabri
authors.affiliation: Research and Development, CSL Limited, Parkville, Victoria, Australia.
authors.forename: Pierre
authors.initials: P
authors.lastname: Scotney
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Australia.|Department of Microbiology, Monash University, Clayton, Australia.
authors.forename: Pantelis
authors.initials: P
authors.lastname: Poumbourios
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Australia heidi.drummer@burnet.edu.au.|Department of Microbiology, Monash University, Clayton, Australia.|Department of Microbiology and Immunology, The University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Australia.
authors.forename: Heidi E
authors.initials: HE
authors.lastname: Drummer
mesh_terms: D000494:Allosteric Regulation; D057134:Antibodies, Neutralizing / Q000737:chemistry; D000914:Antibodies, Viral / Q000737:chemistry; D045744:Cell Line, Tumor; D000939:Epitopes / Q000737:chemistry / Q000276:immunology; D057809:HEK293 Cells; D016174:Hepacivirus / Q000502:physiology*; D022781:Hepatocytes / Q000821:virology; D006801:Humans; D007700:Kinetics; D011485:Protein Binding; D017510:Protein Folding; D054730:Protein Interaction Domains and Motifs; D020836:Protein Structure, Quaternary; D060225:Tetraspanin 28 / Q000737:chemistry; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000502:physiology; D053586:Virus Internalization
chemical_list: D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; C554798:CD81 protein, human; D000939:Epitopes; D060225:Tetraspanin 28; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus
keywords: CD81 receptor; glycoproteins; hepatitis C virus