Background:  Mutations at codon 215 of HIV-1 reverse transcriptase (RT) confer resistance to nucleoside analogs through RT-catalyzed ATP-dependent phosphorolysis.

Methods:  We showed that mutation T215Y is predominant over T215F (respectively, 38.8 vs7.0% of 7312 sequences from a cohort of patients receiving ART in France). Ambiguous mixtures at codon 215 (e.g., TNSY and TSFI) were resolved by cloning and sequencing representative clinical samples. A similar distribution was observed with RT sequences stored in the Stanford HIV Drug Resistance Database. The structural background of these distinct mutational patterns was investigated by molecular modeling of ATP-mutant RT complexes, on the basis of known ATP-protein interactions.

Results:  Accordingly, theoretical profiles TNSY and TSFI were shown to contain almost exclusively T/Y and T/F, respectively. Mutation T215F was preferentially associated with K70R (>71%), D67N (>73%), and K219Q/E/N (>76%), whereas T215Y was associated with M41L (>84%) and L210W (>58%). For the Stanford HIV drug Resistance Database, we found that the aromatic side chain of tyrosine (Y), but not phenylalanine (F), optimally stacked with the adenine ring of ATP. Mutation L210W further stabilized this aromatic pi-pi stacking interaction, increasing the affinity of the T215Y/L210W double mutant for ATP.

Conclusions:  Overall, this study provides a biochemical basis accounting for the evolutionary pathway of T215 mutations in HIV-1 RT, leading to the preferential selection of T215Y vs T215F.