Characterization of Recombinant Integrase of Human Immunodeficiency Virus Type 1 (Isolate Bru)

E. A. Semenova, N. M. Gashnikova, T. V. Il'ina, T. R. Pronyaeva, and A. G. Pokrovsky^*

Institute of Molecular Biology, State Research Center of Virology and Biotechnology “Vector”, Kol'tsovo 630559, Novosibirsk Region, Russia; fax: (3-832) 36-7409; E-mail: pokrovsky@vector.nsc.ru

^* To whom correspondence should be addressed.

Received August 1, 2002

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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 µM) and dependence of integrase activity on Mg²⁺ and Mn²⁺ concentration were studied.

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KEY WORDS: integrase, human immunodeficiency virus type 1, isolate Bru, aurintricarbonic acid

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