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Shinji Harada - One of the best experts on this subject based on the ideXlab platform.

  • isolation of tak 779 resistant hiv 1 from an r5 hiv 1 Gp120 v3 loop library
    Journal of Biological Chemistry, 2005
    Co-Authors: Keisuke Yusa, Yosuke Maeda, Aki Fujioka, Kazuaki Monde, Shinji Harada
    Abstract:

    The human immunodeficiency virus (HIV-1) envelope Glycoprotein (Gp) 120 interacts with CD4 and the CCR5 coreceptor for viral entry. The V3 loop in Gp120 is a crucial region for determining coreceptor usage during viral entry, and a variety of amino acid substitutions has been observed in clinical isolates. To construct an HIV-1 V3 loop library, we chose 10 amino acid positions in the V3 loop and incorporated random combinations (27,648 possibilities) of the amino acid substitutions derived from 31 R5 viruses into the V3 loop of HIV-1(JR-FL) proviral DNA. The constructed HIV-1 library contained 6.6 x 10(6) independent clones containing a set of 0-10 amino acid substitutions in the V3 loop. To address whether restricted steric alteration in the V3 loop could confer resistance to an entry inhibitor, TAK-779, we selected entry inhibitor-resistant HIV-1 by increasing the concentration of TAK-779 from 0.10 to 0.30 microM in PM1-CCR5 cells with high expression of CCR5. The selected viruses at passage 8 contained five amino acid substitutions in the V3 loop without any other mutations in Gp120 and showed 15-fold resistance compared with the parental virus. These results indicated that a certain structure of the V3 loop containing amino acid substitutions derived from 31 R5 viruses can contribute to the acquisition of resistance to entry inhibitors binding to CCR5. Taken together, this type of HIV-1 V3 loop library is useful for isolating and analyzing the specific biological features of HIV-1 with respect to alterations of the V3 loop structure.

Emmanuel Fenouillet - One of the best experts on this subject based on the ideXlab platform.

  • protein disulfide isomerase mediated reduction of two disulfide bonds of hiv envelope Glycoprotein 120 occurs post cxcr4 binding and is required for fusion
    Journal of Biological Chemistry, 2003
    Co-Authors: Rym Barbouche, Raymond Miquelis, Ian M Jones, Emmanuel Fenouillet
    Abstract:

    The human immunodeficiency virus (HIV) envelope (Env) Glycoprotein (Gp) 120 is a highly disulfide-bonded molecule that attaches HIV to the lymphocyte surface receptors CD4 and CXCR4. Conformation changes within Gp120 result from binding and trigger HIV/cell fusion. Inhibition of lymphocyte surface-associated protein-disulfide isomerase (PDI) blocks HIV/cell fusion, suggesting that redox changes within Env are required. Using a sensitive assay based on a thiol reagent, we show that (i) the thiol content of Gp120, either secreted by mammalian cells or bound to a lymphocyte surface enabling CD4 but not CXCR4 binding, was 0.5-1 pmol SH/pmol Gp120 (SH/Gp120), whereas that of Gp120 after its interaction with a surface enabling both CD4 and CXCR4 binding was raised to 4 SH/Gp120; (ii) PDI inhibitors prevented this change; and (iii) Gp120 displaying 2 SH/Gp120 exhibited CD4 but not CXCR4 binding capacity. In addition, PDI inhibition did not impair Gp120 binding to receptors. We conclude that on average two of the nine disulfides of Gp120 are reduced during interaction with the lymphocyte surface after CXCR4 binding prior to fusion and that cell surface PDI catalyzes this process. Disulfide bond restructuring within Env may constitute the molecular basis of the post-receptor binding conformational changes that induce fusion competence.

Richard N Perham - One of the best experts on this subject based on the ideXlab platform.

  • structural mimicry and enhanced immunogenicity of peptide epitopes displayed on filamentous bacteriophage the v3 loop of hiv 1 Gp120
    Journal of Molecular Biology, 1994
    Co-Authors: Fulvia Di Marzo Veronese, Anne E Willis, Cynthia Boyerthompson, Ettore Appella, Richard N Perham
    Abstract:

    Abstract The principal neutralizing determinant of the human immunodeficiency virus type 1 (HIV-1) is an intra-chain disulphide-bridged loop, designated V3, in the third hypervariable region of the surface Glycoprotein Gp 120. Peptide sequences from the V3 loop of Gp120 from HIV-1 strain MN (HIV-1 MN ) were engineered into the N-terminal region of the major coat protein of filamentous bacteriophage fd, leading to their display in multiple copies on the surface of the bacteriophage virion. Peptides displayed in this way were shown to be remarkably effective structural mimics of the natural epitope. They were recognised by human HIV antisera and evoked high titres of antibodies in mice, which cross-reacted with other strains of HIV and were capable of neutralizing the virus. In addition, antibody production could be stimulated by simultaneous inoculation with T-cell epitopes similarly displayed on filamentous bacteriophage. The bacteriophage display system offers a powerful means of studying the immunological recognition of proteins and is a promising vaccine model.

Keisuke Yusa - One of the best experts on this subject based on the ideXlab platform.

  • isolation of tak 779 resistant hiv 1 from an r5 hiv 1 Gp120 v3 loop library
    Journal of Biological Chemistry, 2005
    Co-Authors: Keisuke Yusa, Yosuke Maeda, Aki Fujioka, Kazuaki Monde, Shinji Harada
    Abstract:

    The human immunodeficiency virus (HIV-1) envelope Glycoprotein (Gp) 120 interacts with CD4 and the CCR5 coreceptor for viral entry. The V3 loop in Gp120 is a crucial region for determining coreceptor usage during viral entry, and a variety of amino acid substitutions has been observed in clinical isolates. To construct an HIV-1 V3 loop library, we chose 10 amino acid positions in the V3 loop and incorporated random combinations (27,648 possibilities) of the amino acid substitutions derived from 31 R5 viruses into the V3 loop of HIV-1(JR-FL) proviral DNA. The constructed HIV-1 library contained 6.6 x 10(6) independent clones containing a set of 0-10 amino acid substitutions in the V3 loop. To address whether restricted steric alteration in the V3 loop could confer resistance to an entry inhibitor, TAK-779, we selected entry inhibitor-resistant HIV-1 by increasing the concentration of TAK-779 from 0.10 to 0.30 microM in PM1-CCR5 cells with high expression of CCR5. The selected viruses at passage 8 contained five amino acid substitutions in the V3 loop without any other mutations in Gp120 and showed 15-fold resistance compared with the parental virus. These results indicated that a certain structure of the V3 loop containing amino acid substitutions derived from 31 R5 viruses can contribute to the acquisition of resistance to entry inhibitors binding to CCR5. Taken together, this type of HIV-1 V3 loop library is useful for isolating and analyzing the specific biological features of HIV-1 with respect to alterations of the V3 loop structure.

Victoria R Polonis - One of the best experts on this subject based on the ideXlab platform.

  • safety and immunogenicity of combinations of recombinant subtype e and b human immunodeficiency virus type 1 envelope Glycoprotein 120 vaccines in healthy thai adults
    The Journal of Infectious Diseases, 2003
    Co-Authors: Punnee Pitisuttithum, Mark De Souza, Sorachai Nitayaphan, P Thongcharoen, Chirasak Khamboonruang, Jerome H Kim, Thippawan Chuenchitra, Robin P Garner, Darawan Thapinta, Victoria R Polonis
    Abstract:

    Safety and immunogenicity of 2 recombinant human immunodeficiency virus (HIV) type 1 envelope Glycoprotein (Gp) 120 vaccines derived from SF2 (subtype B) and CM235 (CRF01_AE, Thai E) were evaluated in 370 Thai adults at low risk of HIV infection. Various doses of CM235 (25, 50, or 100 microg) and SF2 (0, 25, or 50 microg) Gp120 were used. Eighty volunteers received placebo. There were no serious adverse events related to vaccination. Binding antibody developed in all vaccine recipients. There was no dose response to CM235 Gp120, but a dose response to Gp120 SF2 was present. Neutralizing antibodies to subtype E HIV-1 NPO3 and subtype B HIV-1 SF2 developed in 84% and 82% of vaccine recipients, respectively. Lymphoproliferative responses were detected in >95% of vaccine recipients. There was no evidence of antigenic interference in HIV-specific humoral or cellular responses. The Gp120 Thai E and SF2 vaccines were safe and immunogenic in combination and could be advanced into phase 3 testing.