Summary information and primary citation

PDB-id
8amd; SNAP-derived features in text and JSON formats; DNAproDB
Class
recombination
Method
cryo-EM (3.9 Å)
Summary
cryo-EM structure of the reca presynaptic filament from s.pneumoniae
Reference
Hertzog M, Perry TN, Dupaigne P, Serres S, Morales V, Soulet AL, Bell JC, Margeat E, Kowalczykowski SC, Le Cam E, Fronzes R, Polard P (2023): "Assembly mechanism and cryoEM structure of RecA recombination nucleofilaments from Streptococcus pneumoniae." Nucleic Acids Res., 51, 2800-2817. doi: 10.1093/nar/gkad080.
Abstract
RecA-mediated homologous recombination (HR) is a key mechanism for genome maintenance and plasticity in bacteria. It proceeds through RecA assembly into a dynamic filament on ssDNA, the presynaptic filament, which mediates DNA homology search and ordered DNA strand exchange. Here, we combined structural, single molecule and biochemical approaches to characterize the ATP-dependent assembly mechanism of the presynaptic filament of RecA from Streptococcus pneumoniae (SpRecA), in comparison to the Escherichia coli RecA (EcRecA) paradigm. EcRecA polymerization on ssDNA is assisted by the Single-Stranded DNA Binding (SSB) protein, which unwinds ssDNA secondary structures that block EcRecA nucleofilament growth. We report by direct microscopic analysis of SpRecA filamentation on ssDNA that neither of the two paralogous pneumococcal SSBs could assist the extension of SpRecA nucleopolymers. Instead, we found that the conserved RadA helicase promotes SpRecA nucleofilamentation in an ATP-dependent manner. This allowed us to solve the atomic structure of such a long native SpRecA nucleopolymer by cryoEM stabilized with ATPĪ³S. It was found to be equivalent to the crystal structure of the EcRecA filament with a marked difference in how RecA mediates nucleotide orientation in the stretched ssDNA. Then, our results show that SpRecA and EcRecA HR activities are different, in correlation with their distinct ATP-dependent ssDNA binding modes.

Cartoon-block schematics in six views (download the tarball)

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