Catalytic efficiency and phenotype of HIV-1 proteases encoding single critical resistance substitutions.
We have shown that a bacteriophage lambda genetic screen system may be useful in predicting the activity and phenotype of HIV-1 protease in the course of viral infection and antiretroviral therapy. This simple and rapid genetic screening system has been used here to characterize HIV-1 proteases encoding single primary resistance substitutions. Except for proteases with amino acid changes at positions 46 and 84, proteases containing single-resistance substitutions displayed a lower catalytic efficiency than the WT enzyme. Single mutants could be identified by their efficiency, demonstrating that modest differences in protease activity can be monitored with this simple assay. Overall, drug susceptibility could be reduced by introduction of single mutations. However, high-level protease inhibitor (PI) resistance was only achieved by multiple mutated proteases. The small but reproducible increase in resistance displayed by single mutants also demonstrated the ability of this genetic screen system for detecting minor reductions in drug susceptibility. These results show that the bacteriophage lambda genetic screen system used here is a useful tool in the analysis of specific contribution of mutations in the HIV protease-coding region or in specific cleavage sites that affect the process of PI resistance.
Fundació irsiCaixa, Laboratori de Retrovirologia, Hospital Universitari Germans Trias i Pujol, 08916, Badalona, Spain.
Marta
M
Cabana
Guerau
G
Fernàndez
Mariona
M
Parera
Bonaventura
B
Clotet
Miguel Angel
MA
Martínez
D019943:Amino Acid Substitution; D001483:Base Sequence; D002384:Catalysis; D017931:DNA Primers; D024882:Drug Resistance, Viral / Q000235:genetics*; D016333:HIV Protease / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D017320:HIV Protease Inhibitors / Q000494:pharmacology*; D015497:HIV-1 / Q000235:genetics; D006801:Humans; D016297:Mutagenesis, Site-Directed; D010641:Phenotype
D017931:DNA Primers; D017320:HIV Protease Inhibitors; D016333:HIV Protease