Publications & Reports

Characterization of novel non-nucleoside reverse transcriptase (RT) inhibitor resistance mutations at residues 132 and 135 in the 51 kDa subunit of HIV-1 RT.

Nissley DV, Radzio J, Ambrose Z, Sheen CW, Hamamouch N, Moore KL, Tachedjian G, Sluis-Cremer N
Basic Research Program, SAIC-Frederick, Inc., NCI-Frederick, Frederick, Maryland 21702, USA.

Abstract

Several rare and novel NNRTI [non-nucleoside reverse transcriptase (RT) inhibitor] resistance mutations were recently detected at codons 132 and 135 in RTs from clinical isolates using the yeast-based chimaeric TyHRT (Ty1/HIV-1 RT) phenotypic assay. Ile132 and Ile135 form part of the beta7-beta8 loop of HIV-1 RT (residues 132-140).

To elucidate the contribution of these residues in RT structure-function and drug resistance, we constructed twelve recombinant enzymes harbouring mutations at codons 132 and 135-140. Several of the mutant enzymes exhibited reduced DNA polymerase activities.

Using the yeast two-hybrid assay for HIV-1 RT dimerization we show that in some instances this decrease in enzyme activity could be attributed to the mutations, in the context of the 51 kDa subunit of HIV-1 RT, disrupting the subunit-subunit interactions of the enzyme.

Drug resistance analyses using purified RT, the TyHRT assay and antiviral assays demonstrated that the I132M mutation conferred high-level resistance (>10-fold) to nevirapine and delavirdine and low-level resistance (approximately 2-3-fold) to efavirenz.

The I135A and I135M mutations also conferred low level NNRTI resistance (approximately 2-fold).

Subunit selective mutagenesis studies again demonstrated that resistance was conferred via the p51 subunit of HIV-1 RT.

Taken together, our results highlight a specific role of residues 132 and 135 in NNRTI resistance and a general role for residues in the beta7-beta8 loop in the stability of HIV-1 RT.

Publication

  • Journal: The Biochemical Journal
  • Published: 15/05/2007
  • Volume: 404
  • Issue: 1
  • Pagination: 151-157

Author

Health Issue