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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
- Source :
- Biochemical Journal. 404:151-157
- Publication Year :
- 2007
- Publisher :
- Portland Press Ltd., 2007.
-
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.
- Subjects :
- Models, Molecular
Protein Conformation
DNA polymerase
Protein subunit
Mutant
Molecular Conformation
Saccharomyces cerevisiae
Drug resistance
medicine.disease_cause
Biochemistry
medicine
Delavirdine
Cloning, Molecular
Molecular Biology
Mutation
biology
Chemistry
Circular Dichroism
Mutagenesis
Cell Biology
Molecular biology
HIV Reverse Transcriptase
Recombinant Proteins
Reverse transcriptase
Protein Subunits
DNA, Viral
HIV-1
biology.protein
RNA, Viral
Reverse Transcriptase Inhibitors
Research Article
medicine.drug
Subjects
Details
- ISSN :
- 14708728 and 02646021
- Volume :
- 404
- Database :
- OpenAIRE
- Journal :
- Biochemical Journal
- Accession number :
- edsair.doi.dedup.....70a9fcb6271bf54ecd81d1a4d7377ff5