Abstract
DNA-restructuring activities of RecQ-family helicases play key roles in genome maintenance. These activities, driven by two tandem RecA-like core domains, are thought to be controlled by accessory DNA-binding elements including the helicase-and-RnaseD-C-terminal (HRDC) domain. The HRDC domain of human Bloom’s syndrome (BLM) helicase was shown to interact with the RecA core, raising the possibility that it may affect the coupling between ATP hydrolysis, translocation along single-stranded (ss)DNA and/or unwinding of double-stranded (ds)DNA. Here, we determined how these activities are affected by the abolition of the ssDNA interaction of the HRDC domain or the deletion of the entire domain in E. coli RecQ helicase. Our data show that the HRDC domain suppresses the rate of DNA-activated ATPase activity in parallel with those of ssDNA translocation and dsDNA unwinding, regardless of the ssDNA binding capability of this domain. The HRDC domain does not affect either the processivity of ssDNA translocation or the tight coupling between the ATPase, translocation, and unwinding activities. Thus, the mechanochemical coupling of E. coli RecQ appears to be independent of HRDC-ssDNA and HRDC-RecA core interactions, which may play roles in more specialized functions of the enzyme.
Highlights
RecQ-family helicases, which are key players in genome maintenance in both bacteria and eukaryotes, perform ssDNA translocation and dsDNA unwinding driven by ATP hydrolysis[1,2,3]
We found that the HRDC domain slows down ssDNA translocation and dsDNA unwinding by suppressing the ATPase activity, but it does not influence the processivity of translocation and the coupling of the ATPase activity to translocation and unwinding
These results suggest that the interaction between the motor core and the HRDC domain is conserved among RecQ helicases and this interaction moderates the rate of ATPase-driven activities, but its effect on mechanochemical coupling may vary among RecQ homologs, according to the differing physiological roles of the enzymes
Summary
RecQ-family helicases, which are key players in genome maintenance in both bacteria and eukaryotes, perform ssDNA translocation and dsDNA unwinding driven by ATP hydrolysis[1,2,3]. In addition we found that, similar to the situation in BLM, the Ec RecQ HRDC domain suppresses the ssDNA-activated ATPase activity of the enzyme[17] These findings suggest a general role for HRDC domains in supporting coupling between ATPase and translocation/unwinding activities through interdomain interactions. We found that the HRDC domain slows down ssDNA translocation and dsDNA unwinding by suppressing the ATPase activity, but it does not influence the processivity of translocation and the coupling of the ATPase activity to translocation and unwinding These results suggest that the interaction between the motor core and the HRDC domain is conserved among RecQ helicases and this interaction moderates the rate of ATPase-driven activities, but its effect on mechanochemical coupling may vary among RecQ homologs, according to the differing physiological roles of the enzymes
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