Characterization of recombinant integrase of human immunodeficiency virus type 1 (isolate Bru).
Integration of the human immunodeficiency virus type 1 (HIV-1) DNA into the human genome requires the virus-encoded integrase protein. The recombinant integrase protein of HIV-1 (isolate Bru) was prepared by constructing a plasmid based on pET-15b encoding the integrase gene. Integrase of HIV-1 was purified using a bacterial expression system (Escherichia coli). The main kinetic parameters of HIV-1 integrase (K(m) = (3.7 +/- 0.2).10(-10) M, k(cat) = (1.2 +/- 0.3).10(-7 )sec(-1)) were determined using an oligonucleotide duplex constructed on the basis of the U5-terminal sequence of proviral HIV-1 DNA as the substrate. Inhibition of integrase by aurintricarbonic acid ([I](50) = 6.3 +/- 0.4 microM) and dependence of integrase activity on Mg2+ and Mn2+ concentration were studied.
Semenova
E A
EA
Institute of Molecular Biology, State Research Center of Virology and Biotechnology Vector, Kol'tsovo 630559, Novosibirsk Region, Russia.
Gashnikova
N M
NM
Il'ina
T V
TV
Pronyaeva
T R
TR
Pokrovsky
A G
AG
D000595:Amino Acid Sequence; D004279:DNA, Viral / Q000378:metabolism; D019427:HIV Integrase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D015497:HIV-1 / Q000201:enzymology* / Q000235:genetics; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism / Q000494:pharmacology; D008345:Manganese / Q000378:metabolism / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*
D004279:DNA, Viral; D011994:Recombinant Proteins; D008345:Manganese; D019427:HIV Integrase; D008274:Magnesium