title: Mutant CD4 molecules with improved binding to HIV envelope protein gp120 selected by phage display.
abstract: Phage-display methodology has been used to select variant CD4 proteins exhibiting increased binding to the surface envelope glycoprotein, gp120, of Human Immunodeficiency Virus Type-1. To facilitate the selection, a library of mutant CD4 proteins was constructed by cloning a PCR-generated error prone population of the first two domains of CD4 into the phagemid expression vector pHEN1. Phage displaying CD4 in functional form were confirmed by Western blot with CD4-specific antibody and by phage ELISA on immobilized gp120. Biopanning of CD4 phage on immobilized gp120 followed by individual characterization identified five clones with increased binding to gp120. All of the selected variants had one or two amino acid substitutions within the V1 domain of CD4, notably at positions 15, 27, 30, 50, and 66 located in the strands surrounding the main binding loop. Variants which exhibited increased binding to recombinant gp120 in vitro were also shown to have an increased capacity for virus neutralization broadly in line with their in vitro binding activity.
authors.affiliation: NERC Institute of Virology, Oxford, United Kingdom.
authors.forename: R
authors.initials: R
authors.lastname: Krykbaev
authors.forename: J
authors.initials: J
authors.lastname: McKeating
authors.forename: I
authors.initials: I
authors.lastname: Jones
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D015704:CD4 Antigens / Q000235:genetics* / Q000378:metabolism; D015723:Gene Library; D015699:HIV Envelope Protein gp120 / Q000378:metabolism*; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D014779:Virus Replication
chemical_list: D015704:CD4 Antigens; D015699:HIV Envelope Protein gp120