Mutations in the thumb allow human immunodeficiency virus type 1 reverse transcriptase to be cleaved by protease in virions

J Virol. 2009 Dec;83(23):12336-44. doi: 10.1128/JVI.00676-09. Epub 2009 Sep 16.

Abstract

Although human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) has been extensively studied, there are still significant questions about the effects of mutations on the maturation and stability of RT. We show here that a significant fraction (>80%) of the single point mutations we generated in the thumb subdomain of HIV-1 (RT) affect the stability of RT in virions. Fragments of the unstable mutant RTs can be detected in Western blots of virion proteins; however, the degree of degradation varies. The titers of the mutants whose virions contain degraded RTs are reduced. Some, but not all, of the unstable RT thumb subdomain mutants we analyzed have a temperature-sensitive phenotype. A preliminary survey of mutations in other subdomains of RT shows that some of these mutations also destabilize RT. The stability of the RT mutants is enhanced by the addition of a protease inhibitor, suggesting that the viral protease plays an important role in the degradation of the mutant RTs. These results confirm and extend earlier reports of mutations that affect the stability of RT in virions. The data suggest that the stability of a mutant RT in virions could be a major factor in determining the virus titer and, by extension, viral fitness, which could affect whether a mutation in RT is acceptable to the virus.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural

MeSH terms

  • HIV Protease / metabolism*
  • HIV Reverse Transcriptase / genetics*
  • HIV Reverse Transcriptase / metabolism*
  • HIV-1 / genetics*
  • Humans
  • Models, Molecular
  • Mutagenesis, Site-Directed
  • Mutant Proteins / genetics
  • Mutant Proteins / metabolism
  • Mutation, Missense*
  • Point Mutation
  • Protein Stability
  • Protein Structure, Tertiary

Substances

  • Mutant Proteins
  • reverse transcriptase, Human immunodeficiency virus 1
  • HIV Reverse Transcriptase
  • HIV Protease
  • p16 protease, Human immunodeficiency virus 1