Interactions with tRNA^Lys induce important structural changes in human immunodeficiency virus reverse transcriptase

Abstract

Retroviral RNA-dependent DNA polymerase (reverse transcriptase or RT) uses the 3'OH end of a cellular tRNA as primer to initiate DNA synthesis. Previous work with avian retrovirus has shown that reverse transcriptase is implicated in the selection of cellular virion-encapsidated tRNAs and has shown that the primer tRNA is positioned on the primer binding site near the 5' end of the viral RNA. These mechanisms support the idea that the retroviral polymerase should form complexes with primer tRNA and the specific encapsidated ones. The genomic sequence of human immunodeficiency virus (HIV) allows the prediction that tRNA LysJ is the natural primer. In this article we show, using the mobility shift assay, that recombinant HIV reverse transcriptase is able to form a complex with bovine tRNALy*. By fluorescence studies and a-chymotrypsin analysis we have observed a modification of the enzyme conformation when reverse transcriptase is bound to the putative primer tRNA. This structural change is specific for tRNALp although the retroviral polymerase is able to interact with other tRNAs.

Forced Selection of a Human Immunodeficiency Virus Type 1 Variant That Uses a Non-Self tRNA Primer for Reverse Transcription: Involvement of Viral RNA Sequences and the Reverse Transcriptase Enzyme

“…One obvious candidate is the reverse transcriptase (RT) polymerase, and a functional analysis of mutant HIV-1 virion particles revealed that RT is indeed involved in the selection and PBS annealing of the tRNA 3 Lys primer (45,57). Biochemical studies have provided additional information on the RT-tRNA 3 Lys complex and its involvement with the RNase H domain (17,54,56,58), but no high-resolution picture has emerged from these studies.…”

A critical role for the TAR element in promoting efficient human immunodeficiency virus type 1 reverse transcription

“…Although the mechanisms responsible for the observed reversions are unknown, the rapid reversion rates suggest that the native HIV-1 PBS is maintained because of stringent selective pressure. Both UV cross-linking and gel retardation analysis indicate that highly specific interactions between HIV-1 reverse transcriptase and its primer tRNA (6) can induce conformational changes in the heterodimeric reverse transcriptase enzyme (48,61). Studies with other retroviruses suggest that RNA sequences outside the PBS are also involved in the control of reverse transcription.…”

Antiviral activity of 4-benzyl pyridinone derivatives as HIV-1 reverse transcriptase inhibitors

“…HIV-1 RT is a very flexible enzyme [20]. Crystal structures have been determined of a complex between the heterodimeric protein and nevirapine [19,21] and other NNRTIs [22], of a ternary complex at 3 Å resolution formed by HIV-1 RT, a of 19 base/18 base double-stranded DNA (primer/template duplex) and a non-inhibitory monoclonal antibody fAb fragment [23].…”