Abstract
Homologous recombination (HR) is a central process of genome biology driven by a conserved recombinase, which catalyses the pairing of single-stranded DNA (ssDNA) with double-stranded DNA to generate a D-loop intermediate. Bacterial RadA is a conserved HR effector acting with RecA recombinase to promote ssDNA integration. The mechanism of this RadA-mediated assistance to RecA is unknown. Here, we report functional and structural analyses of RadA from the human pathogen Streptococcus pneumoniae. RadA is found to facilitate RecA-driven ssDNA recombination over long genomic distances during natural transformation. RadA is revealed as a hexameric DnaB-type helicase, which interacts with RecA to promote orientated unwinding of branched DNA molecules mimicking D-loop boundaries. These findings support a model of DNA branch migration in HR, relying on RecA-mediated loading of RadA hexamers on each strand of the recipient dsDNA in the D-loop, from which they migrate divergently to facilitate incorporation of invading ssDNA.
Highlights
Homologous recombination (HR) is a central process of genome biology driven by a conserved recombinase, which catalyses the pairing of single-stranded DNA with double-stranded DNA to generate a D-loop intermediate
Natural genetic transformation is driven by a dedicated multiprotein machinery, named the transformasome[20], which assembles in the cell membrane and promotes the binding of exogenous double-stranded DNA (dsDNA) on the surface of competent cells, the internalization of one DNA strand and its integration into genomic DNA via RecA-driven homologous recombination (HR) (Fig. 1b)
One explanation of this deficit is that RadA may act to extend single-stranded DNA (ssDNA) incorporation at HR transformation D-loops, a proposal supported by the recent finding that E. coli RadA can promote DNA branch migration in vitro[18]
Summary
Homologous recombination (HR) is a central process of genome biology driven by a conserved recombinase, which catalyses the pairing of single-stranded DNA (ssDNA) with double-stranded DNA to generate a D-loop intermediate. The different HR pathways are driven in their early steps by a common multitasking motor protein (referred to as an HR recombinase) highly conserved in the three domains of life, named RecA in bacteria, Rad[51] and Dmc[1] in eukaryotes and RadA in archaea[4,5] Biochemical analysis of purified E. coli RadA showed that it acts at a 3-way junction (3-J) made by RecA to promote branch migration in an ATP-dependent manner, either with or without RecA18 In this DNA transaction reaction, RadA mimics the DNA strand exchange reaction catalysed by RecA in the 30 direction of the ssDNA paired to the homologous dsDNA template. The helicase migrates divergently from the boundaries of the HR D-loop and is loaded by RecA on both strands of the recipient dsDNA
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
