3os2
PFV target capture complex (TCC) at 3.32 A resolution
Structural highlights
FunctionPOL_FOAMV The aspartyl protease activity mediates proteolytic cleavages of Gag and Pol polyproteins. The reverse transcriptase (RT) activity converts the viral RNA genome into dsDNA in the cytoplasm, shortly after virus entry into the cell (early reverse transcription) or after proviral DNA transcription (late reverse transcription). RT consists of a DNA polymerase activity that can copy either DNA or RNA templates, and a ribonuclease H (RNase H) activity that cleaves the RNA strand of RNA-DNA heteroduplexes in a partially processive 3' to 5' endonucleasic mode. Conversion of viral genomic RNA into dsDNA requires many steps. A tRNA-Lys1,2 binds to the primer-binding site (PBS) situated at the 5'-end of the viral RNA. RT uses the 3' end of the tRNA primer to perform a short round of RNA-dependent minus-strand DNA synthesis. The reading proceeds through the U5 region and ends after the repeated (R) region which is present at both ends of viral RNA. The portion of the RNA-DNA heteroduplex is digested by the RNase H, resulting in a ssDNA product attached to the tRNA primer. This ssDNA/tRNA hybridizes with the identical R region situated at the 3' end of viral RNA. This template exchange, known as minus-strand DNA strong stop transfer, can be either intra- or intermolecular. RT uses the 3' end of this newly synthesized short ssDNA to perform the RNA-dependent minus-strand DNA synthesis of the whole template. RNase H digests the RNA template except for a polypurine tract (PPT) situated at the 5'-end and near the center of the genome. It is not clear if both polymerase and RNase H activities are simultaneous. RNase H probably can proceed both in a polymerase-dependent (RNA cut into small fragments by the same RT performing DNA synthesis) and a polymerase-independent mode (cleavage of remaining RNA fragments by free RTs). Secondly, RT performs DNA-directed plus-strand DNA synthesis using the PPT that has not been removed by RNase H as primer. PPT and tRNA primers are then removed by RNase H. The 3' and 5' ssDNA PBS regions hybridize to form a circular dsDNA intermediate. Strand displacement synthesis by RT to the PBS and PPT ends produces a blunt ended, linear dsDNA copy of the viral genome that includes long terminal repeats (LTRs) at both ends (By similarity). Integrase catalyzes viral DNA integration into the host chromosome, by performing a series of DNA cutting and joining reactions. This enzyme activity takes place after virion entry into a cell and reverse transcription of the RNA genome in dsDNA. The first step in the integration process is 3' processing. This step requires a complex comprising at least the viral genome, matrix protein, and integrase. This complex is called the pre-integration complex (PIC). The integrase protein removes 2 nucleotides from the 3' end of the viral DNA right (U5) end, leaving the left (U3) intact. In the second step, the PIC enters cell nucleus. This process is mediated through the integrase and allows the virus to infect both dividing (nuclear membrane disassembled) and G1/S-arrested cells (active translocation), but with no viral gene expression in the latter. In the third step, termed strand transfer, the integrase protein joins the previously processed 3' ends to the 5' ends of strands of target cellular DNA at the site of integration. It is however not clear how integration then proceeds to resolve the asymmetrical cleavage of viral DNA (By similarity). Publication Abstract from PubMedTo establish productive infection, a retrovirus must insert a DNA replica of its genome into host cell chromosomal DNA. This process is operated by the intasome, a nucleoprotein complex composed of an integrase tetramer (IN) assembled on the viral DNA ends. The intasome engages chromosomal DNA within a target capture complex to carry out strand transfer, irreversibly joining the viral and cellular DNA molecules. Although several intasome/transpososome structures from the DDE(D) recombinase superfamily have been reported, the mechanics of target DNA capture and strand transfer by these enzymes remained unclear. Here we report crystal structures of the intasome from prototype foamy virus in complex with target DNA, elucidating the pre-integration target DNA capture and post-catalytic strand transfer intermediates of the retroviral integration process. The cleft between IN dimers within the intasome accommodates chromosomal DNA in a severely bent conformation, allowing widely spaced IN active sites to access the scissile phosphodiester bonds. Our results resolve the structural basis for retroviral DNA integration and provide a framework for the design of INs with altered target sequences. The mechanism of retroviral integration from X-ray structures of its key intermediates.,Maertens GN, Hare S, Cherepanov P Nature. 2010 Nov 11;468(7321):326-9. PMID:21068843[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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