Mechanistic role of residue Gln151 in error prone DNA synthesis by human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT). Pre-steady state kinetic study of the Q151N HIV-1 RT mutant with increased fidelity

The Journal of Biological Chemistry
K K WeissBaek Kim

Abstract

It has previously been reported that mutations in the Gln(151) residue of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) greatly enhance RT fidelity. In this study, we employed pre-steady state kinetic assays to elucidate the mechanistic role of residue Gln(151) in highly error prone DNA synthesis by HIV-1 RT. Using our Q151N high fidelity mutant, which is structurally altered in its ability to interact with the 3'-OH on the sugar moiety of the incoming deoxynucleotide triphosphate (dNTP), we examined how this change in RT-dNTP interaction affects HIV-1 RT fidelity. First, we found the binding affinity (K(D)) of wild type and Q151N RT proteins to different template/primers to be similar. These results indicate that the Gln(151) residue is not involved in the formation of the binary complex (RT.template/primer) during DNA polymerization. We also found that by changing residue 151 from a Gln-->Asn, the maximum rate of dNTP incorporation (k(pol)) for both correct and incorrect dNTPs was not affected. In contrast, the ability of the Q151N mutant to bind both correct and incorrect dNTPs (K(d)) was diminished. The Q151N mutant was 120-fold less efficient at binding correct dNTP than wild type RT, and its decre...Continue Reading

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Citations

Sep 29, 2004·The Journal of Biological Chemistry·Tracy L DiamondBaek Kim
Nov 5, 2013·AIDS Research and Human Retroviruses·Sarah B LloydWendy R Winnall
Oct 25, 2017·Viral Immunology·Mahmoud Mohammad YaseenMohammad Mahmoud Yaseen
Aug 24, 2006·The Journal of Biological Chemistry·Darwin J OperarioBaek Kim
Nov 19, 2004·The Biochemical Journal·Clara E Cases-González, Luis Menéndez-Arias
Dec 29, 2020·Nucleic Acids Research·Sumit HandaPartho Ghosh

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