title: Design of immunogens that present the crown of the HIV-1 V3 loop in a conformation competent to generate 447-52D-like antibodies.
abstract: gp120 is a subunit of the envelope glycoprotein of HIV-1. The third variable loop region of gp120 (V3 loop) contains multiple immunodominant epitopes and is also functionally important for deciding cell-tropism of the virus. 447-52D is a monoclonal antibody that recognizes the conserved tip of the V3 loop in a beta-turn conformation. This antibody has previously been shown to neutralize diverse strains of the virus. In an attempt to generate an immunogen competent to generate 447-52D-like antibodies, the known epitope of 447-52D was inserted at three different surface loop locations in the small, stable protein Escherichia coli Trx (thioredoxin). At one of the three locations (between residues 74 and 75), the insertion was tolerated, the resulting protein was stable and soluble, and bound 447-52D with an affinity similar to that of intact gp120. Upon immunization, the V3 peptide-inserted Trx scaffold was able to generate anti-V3 antibodies that could compete out 447-52D binding to gp120. Epitope mapping studies demonstrated that these anti-V3 antibodies recognized the same epitope as 447-52D. Although the 447-52D-type antibodies were estimated to be present at concentrations of 50-400 microg/ml of serum, these were not able to effect neutralization of strains like JRFL and BAL but could neutralize the sensitive MN strain. The data suggest that because of the low accessibility of the V3 loop on primary isolates such as JRFL, it will be difficult to elicit a V3-specific, 447-52D-like antibody response to effectively neutralize such isolates.
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560 012, India.
authors.forename: Kausik
authors.initials: K
authors.lastname: Chakraborty
authors.forename: Venuka
authors.initials: V
authors.lastname: Durani
authors.forename: Edward Roshan
authors.initials: ER
authors.lastname: Miranda
authors.forename: Michael
authors.initials: M
authors.lastname: Citron
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Liang
authors.forename: William
authors.initials: W
authors.lastname: Schleif
authors.forename: Joseph G
authors.initials: JG
authors.lastname: Joyce
authors.forename: Raghavan
authors.initials: R
authors.lastname: Varadarajan
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000097:blood / Q000276:immunology*; D000915:Antibody Affinity; D000937:Antigen-Antibody Reactions; D001666:Binding Sites, Antibody / Q000276:immunology; D002942:Circular Dichroism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics; D006168:Guinea Pigs; D015483:HIV Antibodies / Q000097:blood / Q000276:immunology*; D015488:HIV Antigens / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D015497:HIV-1 / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D020349:Surface Plasmon Resonance; D013879:Thioredoxins / Q000737:chemistry / Q000235:genetics
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D029968:Escherichia coli Proteins; D015483:HIV Antibodies; D015488:HIV Antigens; D015699:HIV Envelope Protein gp120; C059762:HIV envelope protein gp120 (305-321); D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D013879:Thioredoxins