Mutational studies in the dNTP binding pocket of human immunodeficiency virus type 1 reverse transcriptase

It was earlier postulated that Gln 91 of HIV-1 RT stabilizes the side chain of Tyr 183 via hydrogen bonding interaction between 0(H) of Y1 83 and CO of Q91 (Harris et al., BIOCHEMISTRY 37: 9630, 1998). In an attempt to understand the function of Gln 91 in the catalytic mechanism, mutants of this residue (Gln91 - -> Ala and Gln91 - ->Asn) were generated and subjected to an in-depth analysis. The efficiency of reverse transcription of natural U5-PBS HIV-1 RNA template was severely impaired by both conservative and non-conservative substitutions from Gln-->Asn and Gln--> Ala, a result similar to that observed with Y-->F substitution at position 183. The major defect seems to be at the substrate binding step as the processivity of these mutants was not affected while the extent of primer utilization correlated with the loss of their polymerase activity. Curiously, these mutant derivatives of Q91 were found to be highly resistant to ddNTPs and exhibited greater stringency in discriminating between correct and incorrect nucleotides. These results suggest possible interaction of Q91 with other residues in the dNTP binding pocket that may be responsible for conferring greater flexibility to the pocket. A model is proposed which suggests that subtle structural changes due to mutation in the region may have influenced the active site of the enzyme that may interfere in the substrate recognition.

Name thesis:degree_name

Master of Science in Biology - (M.S.)

Discipline thesis:degree_discipline

Federated Department of Biological Sciences

Year

2008

Author dc:creator

• Shukla, Smita

Contributors dc:contributor

• Virendra N. Pandey
• Andrew Hill
• G. Miller Jonakait

• Human imunodeficiency virus
• Mutational studies
• Biology

Repository record dc:identifier

https://digitalcommons.njit.edu/theses/362

OAI identifier oai:identifier

oai:digitalcommons.njit.edu:theses-1361