8dwj | pdb_00008dwj
Primase of mutant bacteriophage T4 primosome with single strand DNA/RNA primer hybrid in primer exiting state
Structural highlights
FunctionPRIM_BPT4 Synthesizes short RNA primers for the lagging strand DNA replication (PubMed:2422175, PubMed:3038901, PubMed:11735390, PubMed:22869700). The primase synthesizes short RNA primers on the lagging strand that the polymerase elongates using dNTPs (PubMed:2422175, PubMed:3038901). Recognizes two trinucleotide sequences 5'-GTT-3' and 5'-GCT-3' in vitro, but uses only the first as the priming site in vivo (PubMed:2422175).[HAMAP-Rule:MF_04157][1] [2] [3] [4] Publication Abstract from PubMedThe T4 bacteriophage gp41 helicase and gp61 primase assemble into a primosome complex to couple DNA unwinding with RNA primer synthesis for DNA replication. How a primosome is assembled and how the length of the RNA primer is defined in the T4 bacteriophage, or in any model system, are unclear. Here we report a series of cryo-EM structures of T4 primosome assembly intermediates at resolutions up to 2.7 A. We show that the gp41 helicase is an open spiral in the absence of ssDNA, and ssDNA binding triggers a large-scale scissor-like conformational change that drives the open spiral to a closed ring that activates the helicase. We found that the activation of the gp41 helicase exposes a cryptic hydrophobic primase-binding surface allowing for the recruitment of the gp61 primase. The primase binds the gp41 helicase in a bipartite mode in which the N-terminal Zn-binding domain (ZBD) and the C-terminal RNA polymerase domain (RPD) each contain a helicase-interacting motif (HIM1 and HIM2, respectively) that bind to separate gp41 N-terminal hairpin dimers, leading to the assembly of one primase on the helicase hexamer. Based on two observed primosome conformations - one in a DNA-scanning mode and the other in a post RNA primer-synthesis mode - we suggest that the linker loop between the gp61 ZBD and RPD contributes to the T4 pentaribonucleotide primer. Our study reveals T4 primosome assembly process and sheds light on RNA primer synthesis mechanism. Structural basis of the T4 bacteriophage primosome assembly and primer synthesis.,Feng X, Spiering MM, de Luna Almeida Santos R, Benkovic SJ, Li H bioRxiv. 2023 May 3:2023.05.03.539249. doi: 10.1101/2023.05.03.539249. Preprint. PMID:37205424[5] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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