Analysis of Amino Acids in the β7–β8 Loop of Human Immunodeficiency Virus Type 1 Reverse Transcriptase for their Role in Virus Replication

Abstract

The HIV-1 p51/p66 reverse transcriptase (RT) heterodimer interface comprises, in part, intermolecular interaction of the loop region between β-strands 7 and 8 (β7–β8 loop) in the p51 fingers subdomain with the p66 palm subdomain. In this study, for the first time in the context of infectious HIV-1 particles, we analyzed the contribution of amino acid residues (S134, I135, N136, N137, T139 and P140) in the β7–β8 loop for RT heterodimerization, enzymatic activity, and virus infectivity. Mutating asparagine 136 to alanine (N136A) reduced viral infectivity and enzyme activity dramatically. The N136A mutation appeared to destabilize the RT heterodimer and render both the p66 and p51 subunits susceptible to aberrant cleavage by the viral protease. Subunit-specific mutagenesis demonstrated that the presence of the N136A mutation in the p51 subunit alone was sufficient to cause degradation of RT within the virus particle. Alanine mutation at other residues of the β7–β8 loop did not affect either RT stability or virus infectivity significantly. None of the β7–β8 loop alanine mutations affected the sensitivity of virus to inhibition by NNRTIs. In the context of infectious virions, our results indicate a critical role of the p51 N136 residue within the β7–β8 loop for RT heterodimer stability and function. These findings suggest the interface comprising N136 in p51 and interacting residues in p66 as a possible target for rational drug design.