Cis-Allosteric Regulation of HIV-1 Reverse Transcriptase by Integrase

Abstract

Reverse transcriptase (RT) and integrase (IN) are encoded tandemly in the pol genes of retroviruses. We reported recently that HIV-1 RT and IN need to be supplied as the pol precursor intermediates, in which RT and IN are in fusion form (RTIN) to exert efficient reverse transcription in the context of HIV-1 replication. The mechanism underlying RTIN’s effect, however, remains to be elucidated. In this study, we examined the effect of IN fusion on RT during reverse transcription by an in vitro cell-free assay, using recombinant HIV-1 RTIN (rRTIN). We found that, compared to recombinant RT (rRT), rRTIN generated significantly higher cDNAs under physiological concentrations of dNTPs (less than 10 μM), suggesting increased affinity of RTIN to dNTPs. Importantly, the cleavage of RTIN with HIV-1 protease reduced cDNA levels at a low dose of dNTPs. Similarly, sensitivities against RT inhibitors were significantly altered in RTIN form. Finally, analysis of molecular dynamics simulations of RT and RTIN suggested that IN can influence the structural dynamics of the RT active center and the inhibitor binding pockets in cis. Thus, we demonstrated, for the first time, the cis-allosteric regulatory roles of IN in RT structure and enzymatic activity.

Loxapine inhibits replication of hepatitis A virus in vitro and in vivo by targeting viral protein 2C

“…The resulting model was optimized via energy minimization using the Molecular Operating Environment (MOE) (Chemical Computing Group, Montreal, Quebec, Canada) and an Amber 10: Extended Hu ¨ckel Theory (EHT) force field implemented in MOE, which combines the Amber 10-and EHT-bonded parameters for large-scale energy minimization [31]. The 2C hexamer model was subjected to MD simulations as described previously for viral proteins [32][33][34]. Briefly, the simulations were performed using the pmemd.cuda.MPI module in the Amber 16 program [35] with the ff14SB force field for protein simulation [36].…”