A human IgG1 (b12) specific for the CD4 binding site of HIV-1 neutralizes by inhibiting the virus fusion entry process, but b12 Fab neutralizes by inhibiting a postfusion event.
The human b12 IgG1, specific for the CD4 binding site of the gp120 of HIV-1, was prepared by recombinant DNA technology. It had a high neutralization rate constant (-3.5 x 10(5) M(-1) sec(-1)), although this is about 10-fold less than the values for the best poliovirus or influenza A virus MAbs. The recombinant b12 Fab neutralized well, with about one-tenth of the activity of b12 IgG. The mechanisms by which b12 IgG1 and its Fab neutralize HIV-1 IIIB on C8166 cells have been investigated. Neither inhibited attachment of virus to the target cell as judged by FACS, immunofluorescence, and ELISA data. This was controlled using MAb F105, another human IgG1, that did neutralize by inhibiting attachment under our conditions. The interactions of b12 IgG- and Fab-neutralized virions with target cells were compared with those of nonneutralized virus using a number of different techniques (fluorescence dequenching of R18-labeled virions, immunofluorescence of virion gp41 and p24 antigens, and acquisition of resistance to removal of virions from the cell by protease). These and the inhibition of HIV-1-mediated cell-cell fusion all demonstrated that b12 IgG neutralized by inhibiting the primary fusion-uncoating mechanism. However, the interactions of b12 Fab-neutralized and nonneutralized virions with C8166 cells were indistinguishable. Thus b12 Fab did not inhibit fusion uncoating, and by inference inhibited a stage of infection that occurs after the entry of the virion core into the cytoplasm. It is therefore possible that b12 IgG kills HIV-1 twice over, by fusion-inhibition and by inhibiting the postentry event proposed for the Fab. The mechanism of neutralization of b12 Fab and of other MAbs that neutralize in a similar way and why b12 Fab and IgG neutralize by different mechanisms are discussed.
McInerney
T L
TL
Department of Biological Sciences, University of Warwick, Coventry, United Kingdom.
McLain
L
L
Armstrong
S J
SJ
Dimmock
N J
NJ
D001665:Binding Sites; D015704:CD4 Antigens / Q000276:immunology*; D002459:Cell Fusion; D002460:Cell Line; D019084:Fluorescent Antibody Technique, Indirect; D015483:HIV Antibodies / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000276:immunology*; D006367:HeLa Cells; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology*; D007074:Immunoglobulin G / Q000276:immunology*; D009500:Neutralization Tests; D014357:Trypsin / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014771:Virion
D015704:CD4 Antigens; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D014357:Trypsin