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Research Article
1 December 1987

3'-Azido-3'-deoxythymidine triphosphate as an inhibitor and substrate of purified human immunodeficiency virus reverse transcriptase

Abstract

Reverse transcriptase was purified from human immunodeficiency virus (HIV). It utilized the artificial primer-template poly(rA)-oligo(dT)12-18 more efficiently than activated calf thymus DNA, poly(rI)-oligo(dC)12-18, poly(rC)-oligo(dG)12-18, or poly(rCm)-oligo(dG)12-18. Maximum activity was observed at pH 7.0 to 7.6 in the presence of 5 mM MgCl2 and 100 mM KCl. 3'-Azido-3'-deoxythymidine triphosphate competed with dTTP for binding to HIV reverse transcriptase. Different kinetic constants were obtained with different primer-templates. Km and Ki values of 2.8 and 0.04 microM, respectively, were obtained with poly(rA)-oligo(dT)12-18. The corresponding values were 1.2 and 0.3 microM, respectively, with activated calf thymus DNA and 0.3 and 0.01 microM, respectively, with extracted virus and native template. Inhibition of the host cell DNA polymerases alpha and beta was considerably weaker. The Km and Ki values obtained with activated calf thymus DNA as the primer-template were 2.4 and 230 microM, respectively, for DNA polymerase alpha and 6.0 and 73 microM, respectively, for DNA polymerase beta. 3'-Azido-3'-deoxythymidine triphosphate could also serve as an alternate substrate for HIV reverse transcriptase. The resulting incorporation of 3'-azido-3'-deoxythymidine triphosphate into poly(rA)-oligo(dT)12-18 caused chain termination and premature deceleration of the reaction. The terminated primer could not be elongated when incubated with dTTP and HIV reverse transcriptase.

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Published In

cover image Antimicrobial Agents and Chemotherapy
Antimicrobial Agents and Chemotherapy
Volume 31Number 12December 1987
Pages: 1972 - 1977
PubMed: 2449866

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Published online: 1 December 1987

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Authors

M H St Clair
Department of Virology, Burroughs Wellcome Co., Research Triangle Park, North Carolina 27709.
C A Richards
Department of Virology, Burroughs Wellcome Co., Research Triangle Park, North Carolina 27709.
T Spector
Department of Virology, Burroughs Wellcome Co., Research Triangle Park, North Carolina 27709.
K J Weinhold
Department of Virology, Burroughs Wellcome Co., Research Triangle Park, North Carolina 27709.
W H Miller
Department of Virology, Burroughs Wellcome Co., Research Triangle Park, North Carolina 27709.
A J Langlois
Department of Virology, Burroughs Wellcome Co., Research Triangle Park, North Carolina 27709.
P A Furman
Department of Virology, Burroughs Wellcome Co., Research Triangle Park, North Carolina 27709.

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