Trans-complementation by the RecB nuclease domain of RecBCD enzyme reveals new insight into RecA loading upon χ recognition

Theetha L Pavankumar; C Jason Wong; Yun Ka Wong; Maria Spies; Stephen C Kowalczykowski

doi:10.1093/nar/gkae007

Back

Trans-complementation by the RecB nuclease domain of RecBCD enzyme reveals new insight into RecA loading upon χ recognition

Journal article

Open access

Peer reviewed

Trans-complementation by the RecB nuclease domain of RecBCD enzyme reveals new insight into RecA loading upon χ recognition

Theetha L Pavankumar, C Jason Wong, Yun Ka Wong, Maria Spies and Stephen C Kowalczykowski

Nucleic acids research, Vol.52(5), pp.2578-2589

03/21/2024

DOI: 10.1093/nar/gkae007

PMCID: PMC10954480

PMID: 38261972

Files and links (1)

url

https://doi.org/10.1093/nar/gkae007View

Published (Version of record) Open Access

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.

Details

Title: Subtitle: Trans-complementation by the RecB nuclease domain of RecBCD enzyme reveals new insight into RecA loading upon χ recognition
Creators: Theetha L Pavankumar - University of California, Davis
C Jason Wong - University of California, Davis
Yun Ka Wong - University of California, Davis
Maria Spies - University of Iowa
Stephen C Kowalczykowski - University of California, Davis
Resource Type: Journal article
Publication Details: Nucleic acids research, Vol.52(5), pp.2578-2589
DOI: 10.1093/nar/gkae007
PMID: 38261972
PMCID: PMC10954480
NLM abbreviation: Nucleic Acids Res
ISSN: 0305-1048
eISSN: 1362-4962
Grant note: R35 GM131900 / NIH HHS
Language: English
Electronic publication date: 01/23/2024
Date published: 03/21/2024
Academic Unit: Radiation Oncology; Biochemistry and Molecular Biology
Record Identifier: 9984548850502771

Metrics

6 Record Views

5 Times Cited - Web of Science