I. Olivares et al., Second-site reversion of a human immunodeficiency virus type 1 reverse transcriptase mutant that restores enzyme function and replication capacity, J VIROLOGY, 73(8), 1999, pp. 6293-6298
Nonconservative substitutions for Tyr-115 in the reverse transcriptase (RT)
of human immunodeficiency virus type 1 (HIV-1) lead to enzymes displaying
lower affinity for deoxynucleoside triphosphates (dNTPs) (A. M. Martin-Hern
andez, E. Domingo, and L. Menendez-Arias, EMBO J. 15:4434-4442, 1996). Seve
ral mutations at this position (Y115W, Y115L, Y115A, and Y115D) were introd
uced in an infectious HIV-1 clone, and the replicative capacity of the muta
nt viruses was monitored. Y115W was the only mutant able to replicate in MT
-4 cells, albeit very poorly. Nucleotide sequence analysis of the progeny v
irus recovered from supernatants of four independent transfection experimen
ts showed that the Y115W mutation was maintained. However, in all cases an
additional substitution in the primer grip of the RT (M230I) emerged when t
he virus increased its replication capacity. Using recombinant HIV-1 RT, we
demonstrate that M230I mitigates the polymerase activity defect of the Y11
5W mutant, by increasing the dNTP binding affinity of the enzyme. The secon
d-site suppressor effects observed were mediated by mutations in the 66-kDa
subunit of the RT, as demonstrated with chimeric heterodimers. Examination
of available crystal structures of HIV-1 RT suggests a possible mechanism
for restoration of enzyme activity by the second-site revertant.