Abstract:
Reverse transcription of the HIV-1 RNA genome into double-stranded DNA is a central step in viral infection ¹ and a common target of antiretroviral drugs ² . The reaction is catalysed by viral reverse transcriptase (RT)^3,4 that is packaged in an infectious virion with two copies of viral genomic RNA ⁵ each bound to host lysine 3 transfer RNA (tRNA^Lys₃), which acts as a primer for initiation of reverse transcription^6,7. Upon viral entry into cells, initiation is slow and non-processive compared to elongation^8,9. Despite extensive efforts, the structural basis of RT function during initiation has remained a mystery. Here we use cryo-electron microscopy to determine a three-dimensional structure of an HIV-1 RT initiation complex. In our structure, RT is in an inactive polymerase conformation with open fingers and thumb and with the nucleic acid primer-template complex shifted away from the active site. The primer binding site (PBS) helix formed between tRNA^Lys₃ and HIV-1 RNA lies in the cleft of RT and is extended by additional pairing interactions. The 5' end of the tRNA refolds and stacks on the PBS to create a long helical structure, while the remaining viral RNA forms two helical stems positioned above the RT active site, with a linker that connects these helices to the RNase H region of the PBS. Our results illustrate how RNA structure in the initiation complex alters RT conformation to decrease activity, highlighting a potential target for drug action.