title: Identification of amino acid residues in CD81 critical for interaction with hepatitis C virus envelope glycoprotein E2.
abstract: Human CD81 has been previously identified as the putative receptor for the hepatitis C virus envelope glycoprotein E2. The large extracellular loop (LEL) of human CD81 differs in four amino acid residues from that of the African green monkey (AGM), which does not bind E2. We mutated each of the four positions in human CD81 to the corresponding AGM residues and expressed them as soluble fusion LEL proteins in bacteria or as complete membrane proteins in mammalian cells. We found human amino acid 186 to be critical for the interaction with the viral envelope glycoprotein. This residue was also important for binding of certain anti-CD81 monoclonal antibodies. Mutating residues 188 and 196 did not affect E2 or antibody binding. Interestingly, mutation of residue 163 increased both E2 and antibody binding, suggesting that this amino acid contributes to the tertiary structure of CD81 and its ligand-binding ability. These observations have implications for the design of soluble high-affinity molecules that could target the CD81-E2 interaction site(s).
authors.affiliation: Department of Molecular Biology, University of Sheffield, Sheffield S10 2UH, United Kingdom.
authors.forename: A
authors.initials: A
authors.lastname: Higginbottom
authors.forename: E R
authors.initials: ER
authors.lastname: Quinn
authors.forename: C C
authors.initials: CC
authors.lastname: Kuo
authors.forename: M
authors.initials: M
authors.lastname: Flint
authors.forename: L H
authors.initials: LH
authors.lastname: Wilson
authors.forename: E
authors.initials: E
authors.lastname: Bianchi
authors.forename: A
authors.initials: A
authors.lastname: Nicosia
authors.forename: P N
authors.initials: PN
authors.lastname: Monk
authors.forename: J A
authors.initials: JA
authors.lastname: McKeating
authors.forename: S
authors.initials: S
authors.lastname: Levy
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000915:Antibody Affinity; D000936:Antigen-Antibody Complex; D015703:Antigens, CD / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D001665:Binding Sites; D002460:Cell Line; D002522:Chlorocebus aethiops; D016174:Hepacivirus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D017354:Point Mutation; D011487:Protein Conformation; D011956:Receptors, Cell Surface / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D060225:Tetraspanin 28; D013861:Thiocyanates / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D015703:Antigens, CD; C554798:CD81 protein, human; D008565:Membrane Proteins; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; D060225:Tetraspanin 28; D013861:Thiocyanates; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus; C026976:thiocyanic acid