title: Germlining of the HIV-1 broadly neutralizing antibody domain m36.
abstract: Engineered antibody domains (eAds) have emerged as a novel class of HIV-1 inhibitors and are currently under preclinical testing as promising drug candidates for prevention and therapy of HIV-1 infection. Reverse mutation of antibodies to germline sequences (germlining) could not only identify less mutated variants with lower probability of immunogenicity and other improved properties but also help elucidate their mechanisms of action. In this study, we sequentially reverted the framework (FRs) and complementary determining regions (CDRs) of m36, a human antibody heavy chain variable domain-based eAd targeting the coreceptor binding site of the viral envelope glycoprotein gp120, back to germline sequences. Two types of amino acid mutations and one region in the antibody V segment were identified that are critical for HIV-1 neutralization. These include four mutations to acidic acid residues distributed in the CDR1 and CDR2, two mutations to hydrophobic residues in the FR3 and CDR3, and partial FR2 and FR3 sequences flanking the CDR2 that are derived from a different gene family. An m36 variant with all five mutations in the FRs reverted back to germline showed slightly increased neutralizing activity against two HIV-1 isolates tested. Another variant with seven of twelve mutations in the V segment reverted retained potency within threefold of that of the mature antibody. These results, together with an analysis of m36-gp120-CD4 docking structures, could have implications for the further development of m36 and elucidation of its mechanism of potent and broad HIV-1 neutralization.
authors: Protein Interactions Section, Laboratory of Experimental Immunology, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA. Electronic address: chenw3@mail.nih.gov.
authors: Weizao
authors: W
authors: Chen
authors: Protein Interactions Section, Laboratory of Experimental Immunology, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors: Wei
authors: W
authors: Li
authors: Protein Interactions Section, Laboratory of Experimental Immunology, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors: Tianlei
authors: T
authors: Ying
authors: Protein Interactions Section, Laboratory of Experimental Immunology, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA; The Geneva Foundation, WA 98402, USA.
authors: Yanping
authors: Y
authors: Wang
authors: Protein Interactions Section, Laboratory of Experimental Immunology, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors: Yang
authors: Y
authors: Feng
authors: Protein Interactions Section, Laboratory of Experimental Immunology, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors: Dimiter S
authors: DS
authors: Dimitrov
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing / Q000737:chemistry* / Q000276:immunology*; D001483:Base Sequence; D001666:Binding Sites, Antibody; D022801:Complementarity Determining Regions; D015483:HIV Antibodies / Q000737:chemistry* / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000276:immunology* / Q000378:metabolism; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000276:immunology; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D022801:Complementarity Determining Regions; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins
keywords: Antibody domain; Germlining; HIV-1; Mutation; Neutralization