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Int. J. Mol. Sci. 2018, 19(10), 3049; https://doi.org/10.3390/ijms19103049

Movement of the RecG Motor Domain upon DNA Binding Is Required for Efficient Fork Reversal

1
Department of Biological Sciences, Vanderbilt University, Nashville, TN 37232, USA
2
Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN 37232, USA
*
Author to whom correspondence should be addressed.
Received: 1 September 2018 / Revised: 29 September 2018 / Accepted: 4 October 2018 / Published: 6 October 2018
(This article belongs to the Special Issue DNA Replication Stress)
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Abstract

RecG catalyzes reversal of stalled replication forks in response to replication stress in bacteria. The protein contains a fork recognition (“wedge”) domain that binds branched DNA and a superfamily II (SF2) ATPase motor that drives translocation on double-stranded (ds)DNA. The mechanism by which the wedge and motor domains collaborate to catalyze fork reversal in RecG and analogous eukaryotic fork remodelers is unknown. Here, we used electron paramagnetic resonance (EPR) spectroscopy to probe conformational changes between the wedge and ATPase domains in response to fork DNA binding by Thermotoga maritima RecG. Upon binding DNA, the ATPase-C lobe moves away from both the wedge and ATPase-N domains. This conformational change is consistent with a model of RecG fully engaged with a DNA fork substrate constructed from a crystal structure of RecG bound to a DNA junction together with recent cryo-electron microscopy (EM) structures of chromatin remodelers in complex with dsDNA. We show by mutational analysis that a conserved loop within the translocation in RecG (TRG) motif that was unstructured in the RecG crystal structure is essential for fork reversal and DNA-dependent conformational changes. Together, this work helps provide a more coherent model of fork binding and remodeling by RecG and related eukaryotic enzymes. View Full-Text
Keywords: DNA replication; DNA repair; DNA damage response; DNA translocation; DNA helicase; superfamily 2 ATPase; replication restart; fork reversal; fork regression; chromatin remodeler DNA replication; DNA repair; DNA damage response; DNA translocation; DNA helicase; superfamily 2 ATPase; replication restart; fork reversal; fork regression; chromatin remodeler
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).

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Warren, G.M.; Stein, R.A.; Mchaourab, H.S.; Eichman, B.F. Movement of the RecG Motor Domain upon DNA Binding Is Required for Efficient Fork Reversal. Int. J. Mol. Sci. 2018, 19, 3049.

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