title: The stoichiometry of trimeric SIV glycoprotein interaction with CD4 differs from that of anti-envelope antibody Fab fragments.
abstract: Human and simian immunodeficiency viruses infect host lymphoid cells by binding CD4 molecules via their gp160 envelope glycoproteins. Biochemical studies on recombinant SIVmac32H (pJ5) envelope ectodomain gp140 precursor protein show that the envelope is a trimer. Using size exclusion chromatography, quantitative amino acid analysis, analytical ultracentrifugation, and CD4-based competition assay, we demonstrate that the stoichiometry of CD4 receptor-oligomeric envelope interaction is 1:1. By contrast, Fab fragments of both neutralizing and non-neutralizing monoclonal antibodies bind at a 3:1 ratio. Thus, despite displaying equivalent CD4 binding sites on each of the three gp140 protomers within an uncleaved trimer, only one site binds the soluble 4-domain human CD4 extracellular segment. The anti-cooperativity and the faster k(off) of gp140 trimer:CD4 versus gp120 monomer:CD4 interaction suggest that CD4-induced conformational change is impeded in the intact envelope. The implications of these findings for immunity against human immunodeficiency virus and simian immunodeficiency virus are discussed.
authors.affiliation: Laboratory of Immunobiology, Dana-Farber Cancer Institute and Department of Medicine, Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: M
authors.initials: M
authors.lastname: Kim
authors.forename: B
authors.initials: B
authors.lastname: Chen
authors.forename: R E
authors.initials: RE
authors.lastname: Hussey
authors.forename: Y
authors.initials: Y
authors.lastname: Chishti
authors.forename: D
authors.initials: D
authors.lastname: Montefiori
authors.forename: J A
authors.initials: JA
authors.lastname: Hoxie
authors.forename: O
authors.initials: O
authors.lastname: Byron
authors.forename: G
authors.initials: G
authors.lastname: Campbell
authors.forename: S C
authors.initials: SC
authors.lastname: Harrison
authors.forename: E L
authors.initials: EL
authors.lastname: Reinherz
mesh_terms: D000596:Amino Acids / Q000737:chemistry; D001665:Binding Sites; D015153:Blotting, Western; D015704:CD4 Antigens / Q000737:chemistry* / Q000378:metabolism; D002845:Chromatography; D019281:Dimerization; D015686:Gene Products, env / Q000737:chemistry*; D006023:Glycoproteins / Q000737:chemistry*; D006678:HIV / Q000378:metabolism; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D007700:Kinetics; D008024:Ligands; D015233:Models, Statistical; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D015682:Retroviridae Proteins, Oncogenic / Q000737:chemistry*; D020349:Surface Plasmon Resonance; D013997:Time Factors; D014461:Ultracentrifugation; D014760:Viral Fusion Proteins / Q000737:chemistry*
chemical_list: D000596:Amino Acids; D015704:CD4 Antigens; C033904:GP 140; D015686:Gene Products, env; D006023:Glycoproteins; D007140:Immunoglobulin Fab Fragments; D008024:Ligands; D015682:Retroviridae Proteins, Oncogenic; D014760:Viral Fusion Proteins; C059580:transmembrane protein, Simian immunodeficiency virus