Dineshkumar Manvar - The Glutamine Side Chain at Position 91 on the β5a−β5b Loop of Human Immunodeficiency Virus Type 1 Reverse Transcriptase Is Required for Stabilizing the dNTP Binding Pocket

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      Publication Details (including relevant citation   information):

      Biochemistry, 2011, 50 (37), 8067-8077


      Earlier, we postulated that Gln91 of human immunodeficiency virus   type 1 reverse transcriptase (HIV-1 RT) stabilizes the side chain   of Tyr183 via hydrogen bonding interaction between O(H) of Tyr183   and CO of Q91 [Harris, D., et al. (1998) Biochemistry   37, 9630–9640]. To test this hypothesis, we generated mutant   derivatives of Gln91 and analyzed their biochemical properties.   The efficiency of reverse transcription was severely impaired by   nonconservative substitution of Gln with Ala, while conservative   substitution of Gln with Asn resulted in an approximately 70%   loss of activity, a value similar to that observed with the Y183F   mutation. The loss of polymerase activity from both Q91A and Q91N   was significantly improved by a Met to Val substitution at   position 184. Curiously, the Q91N mutant exhibited stringency in   discriminating between correct and incorrect nucleotides,   suggesting its possible interaction with residues influencing the   flexibility of the dNTP binding pocket. In contrast, both double   mutants, Q91A/M184V and Q91N/M184V, are found to be as error   prone as the wild-type enzyme. We propose a model that suggests   that subtle structural changes in the region due to mutation at   position 91 may influence the stability of the side chain of   Tyr183 in the catalytic YMDD motif of the enzyme, thus altering   the active site geometry that may interfere in substrate   recognition.

      Address (URL): http://pubs.acs.org/doi/full/10.1021/bi200815e