Characterization of the p68/p58 Heterodimer of Human Immunodeficiency Virus Type 2 Reverse Transcriptase
| Autor: | Naisheng Fan, S K Sharma, W. G. Tarpley, S M Poppe, K B Rank |
|---|---|
| Rok vydání: | 1996 |
| Předmět: |
Poly T
Cations Divalent Protein Conformation Protein subunit Molecular Sequence Data Ribonuclease H In Vitro Techniques RNA hydrolysis Biochemistry Substrate Specificity law.invention chemistry.chemical_compound HIV Protease law Aspartic Acid Endopeptidases Humans Thymine Nucleotides RNase H DNA Primers Base Sequence biology Chemistry virus diseases RNA RNA-Directed DNA Polymerase Genes gag Molecular biology HIV Reverse Transcriptase Recombinant Proteins Reverse transcriptase Kinetics Terminator (genetics) DNA Viral HIV-2 biology.protein Recombinant DNA Reverse Transcriptase Inhibitors Poly A Protein Processing Post-Translational DNA Dideoxynucleotides |
| Zdroj: | Biochemistry. 35:1911-1917 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/bi9516440 |
| Popis: | Recently we demonstrated that the p58 subunit of p68/p58 HIV-2 reverse transcriptase (RT) heterodimer, produced by processing of p68/p68 homodimer with recombinant HIV-2 protease, terminates at Met484 [Fan, N., et al. (1995) J. Biol. Chem. 270, 13573-13579]. Here we describe purification and characterization of the p68/p58 heterodimer of recombinant HIV-2 RT. It exhibited both RT and RNase H activities, obeyed Michaelis-Menten kinetics, and was competitively inhibited by the DNA chain terminator ddTTP (Ki[app] = 305 +/- 20 nM). The HIV-2 RT-associated RNase H exhibited a marked preference for RNA hydrolysis from a HIV-1 gag-based heteropolymeric RNA/DNA hybrid in the presence of either Mg2+ or Mn2+, compared to the [3H]poly(rA).poly(dT) or [3H]poly(rG).poly(dC) homopolymeric substrates. Relative to HIV-1 RT, the RNase H activity of HIV-2 RT was only 5% toward the [3H]poly(rA).poly(dT) in the presence of Mg2+. The size distribution of products generated from [3H]poly(rA).poly(dT) by HIV-2 RT-associated RNase H was markedly distinct from that of HIV-1 RT in the presence of Mg2+ or Mn2+. The p68/p58 HIV-2 RT heterodimer, produced by specific cleavage using HIV-2 protease, should be useful for inhibition and biophysical studies aimed at discovering and designing drugs directed toward HIV-2. |
| Databáze: | OpenAIRE |
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