Interaction of the reverse transcriptase of human immunodeficiency virus type 1 with DNA.
During DNA synthesis, the binding of human immunodeficiency virus (HIV) reverse transcriptase (RT) to the template-primer precedes its binding to nucleotide triphosphates. The interaction of oligonucleotide DNA with HIV-1 RT was investigated by using a gel retardation assay. Both homodimeric (p66/p66) and heterodimeric (p66/p51) isoforms of HIV-1 RT were capable of binding the DNA oligomers. Thus, all further studies on the interaction of HIV-1 RT with DNA were done with heterodimeric RT. We have studied the conditions for optimal binding. The formation of the RT-DNA complex was primer-independent, and the extent of DNA binding was indistinguishable for both single-stranded and double-stranded DNA (either blunt-ended or recessed). The DNA binding activity of the RT was found to be dependent on oligonucleotide length. HIV-1 RT binds DNA with no apparent sequence specificity. Hence, this enzyme belongs to the sequence nonspecific DNA binding proteins. The interaction was found to be independent of DNA synthesis. The formation of the RT-DNA complex was not influenced by the presence of either template-complementary or noncomplementary dNTPs, indicating that neither DNA polymerization nor binding of the RT to the dNTP affects the stability of the complex. The gel retardation assay was utilized to examine also the effect of various HIV-1 RT inhibitors (i.e., AZT-TP, ddTTP, TIBO, and 3,5,8-trihydroxy-4-quinolone) on the enzyme-DNA interaction. The results indicate differences in the modes of action of these compounds.(ABSTRACT TRUNCATED AT 250 WORDS)
Bakhanashvili
M
M
Department of Cell Biology and Histology, Sackler School of Medicine, Tel Aviv University, Israel.
Hizi
A
A
D000998:Antiviral Agents / Q000494:pharmacology; D001345:Autoradiography; D001483:Base Sequence; D001569:Benzodiazepines / Q000494:pharmacology; D001667:Binding, Competitive; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000187:drug effects* / Q000378:metabolism; D054306:Dideoxynucleotides; D004591:Electrophoresis, Polyacrylamide Gel; D054303:HIV Reverse Transcriptase; D015497:HIV-1 / Q000235:genetics*; D007093:Imidazoles / Q000494:pharmacology; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000378:metabolism*; D015363:Quinolones / Q000494:pharmacology; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018894:Reverse Transcriptase Inhibitors; D013942:Thymine Nucleotides / Q000494:pharmacology; D015215:Zidovudine / Q000031:analogs & derivatives / Q000494:pharmacology
C086339:3,5,8-trihydroxy-4-quinolone; D000998:Antiviral Agents; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D054306:Dideoxynucleotides; D007093:Imidazoles; D009838:Oligodeoxyribonucleotides; D015363:Quinolones; D011994:Recombinant Proteins; D018894:Reverse Transcriptase Inhibitors; D013942:Thymine Nucleotides; D001569:Benzodiazepines; D015215:Zidovudine; C067413:R-82913; C052873:zidovudine triphosphate; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase; C017216:2',3'-dideoxythymidine triphosphate