title: Structural basis of clade-specific HIV-1 neutralization by humanized anti-V3 monoclonal antibody KD-247.
abstract: Humanized monoclonal antibody KD-247 targets the Gly(312)-Pro(313)-Gly(314)-Arg(315) arch of the third hypervariable (V3) loop of the HIV-1 surface glycoprotein. It potently neutralizes many HIV-1 clade B isolates, but not of other clades. To understand the molecular basis of this specificity, we solved a high-resolution (1.55 Å) crystal structure of the KD-247 antigen binding fragment and examined the potential interactions with various V3 loop targets. Unlike most antibodies, KD-247 appears to interact with its target primarily through light chain residues. Several of these interactions involve Arg(315) of the V3 loop. To evaluate the role of light chain residues in the recognition of the V3 loop, we generated 20 variants of KD-247 single-chain variable fragments with mutations in the antigen-binding site. Purified proteins were assessed for V3 loop binding using AlphaScreen technology and for HIV-1 neutralization. Our data revealed that recognition of the clade-specificity defining residue Arg(315) of the V3 loop is based on a network of interactions that involve Tyr(L32), Tyr(L92), and Asn(L27d) that directly interact with Arg(315), thus elucidating the molecular interactions of KD-247 with its V3 loop target.
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Karen A
authors.initials: KA
authors.lastname: Kirby
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Yee Tsuey
authors.initials: YT
authors.lastname: Ong
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Atsuko
authors.initials: A
authors.lastname: Hachiya
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Thomas G
authors.initials: TG
authors.lastname: Laughlin
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Leslie A
authors.initials: LA
authors.lastname: Chiang
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Yun
authors.initials: Y
authors.lastname: Pan
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Moran
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Bruno
authors.initials: B
authors.lastname: Marchand
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine.
authors.forename: Kamalendra
authors.initials: K
authors.lastname: Singh
authors.affiliation: Structural Biology Core, and.
authors.forename: Fabio
authors.initials: F
authors.lastname: Gallazzi
authors.affiliation: Department of Biochemistry, University of Missouri, Columbia, Missouri, USA;
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Quinn
authors.affiliation: AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan;
authors.forename: Kazuhisa
authors.initials: K
authors.lastname: Yoshimura
authors.affiliation: The Chemo-Sero-Therapeutic Research Institute (Kaketsuken), Kyokushi, Kikuchi, Kumamoto, Japan; and.
authors.forename: Toshio
authors.initials: T
authors.lastname: Murakami
authors.affiliation: Division of Clinical Retrovirology and Infectious Diseases, Center for AIDS Research, Kumamoto University, Kumamoto, Japan.
authors.forename: Shuzo
authors.initials: S
authors.lastname: Matsushita
authors.affiliation: Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, School of Medicine, Department of Biochemistry, University of Missouri, Columbia, Missouri, USA; sarafianoss@missouri.edu.
authors.forename: Stefan G
authors.initials: SG
authors.lastname: Sarafianos
mesh_terms: D000595:Amino Acid Sequence; D061067:Antibodies, Monoclonal, Humanized / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D057134:Antibodies, Neutralizing / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D000936:Antigen-Antibody Complex / Q000737:chemistry / Q000235:genetics; D001666:Binding Sites, Antibody / Q000235:genetics; D018360:Crystallography, X-Ray; D015483:HIV Antibodies / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D015497:HIV-1 / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009500:Neutralization Tests; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D054730:Protein Interaction Domains and Motifs; D017386:Sequence Homology, Amino Acid; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D055672:Static Electricity
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D057134:Antibodies, Neutralizing; D000936:Antigen-Antibody Complex; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; C059762:HIV envelope protein gp120 (305-321); D010446:Peptide Fragments; D057127:Single-Chain Antibodies
keywords: HIV; crystal structure; entry; single-chain variable fragment