Human recombinant antibody fragments neutralizing human immunodeficiency virus type 1 reverse transcriptase provide an experimental basis for the structural classification of the DNA polymerase family.
We describe in this paper the binding and biochemical properties of two human antibody fragments directed against the human immunodeficiency virus type 1 reverse transcriptase (RT). These fragments were isolated from a synthetic combinatorial library of human Fab antibody fragments displayed on the surface of filamentous phage. The antibody fragments were selected by using recombinant heterodimeric human immunodeficiency virus type 1 RT purified from insect cells as a solid-phase selector. This procedure led to the isolation of two antibody fragments that completely neutralize the RNA-dependent DNA polymerase activity of RT at nanomolar concentrations. Both antibody fragments bind only to the enzymatically active form of the RT. The inhibitory activity of the anti-RT antibody fragments is competitive with respect to the template primer. The antibody fragments also neutralize the activities of RTs from avian and murine retroviruses and of DNA polymerases of prokaryotic origin as well as human DNA polymerase alpha. Thus, the antibody fragments selected and characterized in this study appear to recognize a structural fold that is common to the different DNA polymerases and necessary for their activity. The results provide an immunological experimental basis for a purely structural and evolutionary classification of the polymerase family.
Gargano
N
N
International School for Advanced Studies, Trieste, Italy.
Biocca
S
S
Bradbury
A
A
Cattaneo
A
A
D000818:Animals; D009189:Avian Myeloblastosis Virus / Q000201:enzymology; D002460:Cell Line; D003090:Coliphages; D015723:Gene Library; D015483:HIV Antibodies / Q000276:immunology*; D054303:HIV Reverse Transcriptase / Q000235:genetics / Q000276:immunology*; D015497:HIV-1 / Q000201:enzymology*; D006801:Humans; D051379:Mice; D008979:Moloney murine leukemia virus / Q000201:enzymology; D009500:Neutralization Tests; D012194:RNA-Directed DNA Polymerase / Q000145:classification / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D018411:Spodoptera / Q000166:cytology
D015483:HIV Antibodies; D011993:Recombinant Fusion Proteins; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase