Architecture of an HIV-1 reverse transcriptase initiation complex

Reverse transcription of the HIV-1 RNA genome into doublestranded DNA is a central step in viral infection1 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(3)(Lys)), 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(3)(Lys) 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.