Trans-complementation by the RecB nuclease domain of RecBCD enzyme reveals new insight into RecA loading upon χ recognition.
Nucleic Acids Res
; 52(5): 2578-2589, 2024 Mar 21.
Article
em En
| MEDLINE
| ID: mdl-38261972
ABSTRACT
The loading of RecA onto ssDNA by RecBCD is an essential step of RecBCD-mediated homologous recombination. RecBCD facilitates RecA-loading onto ssDNA in a χ-dependent manner via its RecB nuclease domain (RecBn). Before recognition of χ, RecBn is sequestered through interactions with RecBCD. It was proposed that upon χ-recognition, RecBn undocks, allowing RecBn to swing out via a contiguous 70 amino acid linker to reveal the RecA-loading surface, and then recruit and load RecA onto ssDNA. We tested this hypothesis by examining the interactions between RecBn (RecB928-1180) and truncated RecBCD (RecB1-927CD) lacking the nuclease domain. The reconstituted complex of RecB1-927CD and RecBn is functional in vitro and in vivo. Our results indicate that despite being covalently severed from RecB1-927CD, RecBn can still load RecA onto ssDNA, establishing that RecBn does not function while only remaining tethered to the RecBCD complex via the linker. Instead, RecBCD undergoes a χ-induced intramolecular rearrangement to reveal the RecA-loading surface.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Recombinases Rec A
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Proteínas de Escherichia coli
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Exodesoxirribonuclease V
Idioma:
En
Revista:
Nucleic Acids Res
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Nucleic acids res
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Nucleic acids research
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
Estados Unidos
País de publicação:
Reino Unido