Single-stranded DNA translocation of E. coli UvrD monomer is tightly coupled to ATP hydrolysis.

Journal of Molecular Biology
Eric J TomkoT M Lohman

Abstract

Escherichia coli UvrD is an SF1A (superfamily 1 type A) helicase/translocase that functions in several DNA repair pathways. A UvrD monomer is a rapid and processive single-stranded DNA (ssDNA) translocase but is unable to unwind DNA processively in vitro. Based on data at saturating ATP (500 μM), we proposed a nonuniform stepping mechanism in which a UvrD monomer translocates with biased (3' to 5') directionality while hydrolyzing 1 ATP per DNA base translocated, but with a kinetic step size of 4-5 nt/step, suggesting that a pause occurs every 4-5 nt translocated. To further test this mechanism, we examined UvrD translocation over a range of lower ATP concentrations (10-500 μM ATP), using transient kinetic approaches. We find a constant ATP coupling stoichiometry of ∼1 ATP/DNA base translocated even at the lowest ATP concentration examined (10 μM), indicating that ATP hydrolysis is tightly coupled to forward translocation of a UvrD monomer along ssDNA with little slippage or futile ATP hydrolysis during translocation. The translocation kinetic step size remains constant at 4-5 nt/step down to 50 μM ATP but increases to ∼7 nt/step at 10 μM ATP. These results suggest that UvrD pauses more frequently during translocation at low AT...Continue Reading

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Citations

May 23, 2013·Nature Communications·Kyung Suk LeeTaekjip Ha
Apr 1, 2015·Nucleic Acids Research·Vessela PetrovaMichael M Cox
Jul 17, 2016·Journal of Molecular Biology·Michael J SimonTimothy M Lohman
Nov 1, 2017·Proceedings of the National Academy of Sciences of the United States of America·Binh NguyenTimothy M Lohman
May 7, 2014·The Journal of Biological Chemistry·Matthew J MeinersSteven W Matson
Nov 5, 2014·Biomolecular Concepts·Koan Briggs, Christopher J Fischer
Sep 17, 2017·The Biochemical Journal·Kaustubh ShuklaGanesh Nagaraju
May 18, 2016·Proceedings of the National Academy of Sciences of the United States of America·Joshua E SokoloskiTimothy M Lohman
Aug 1, 2019·Proceedings of the National Academy of Sciences of the United States of America·Yerdos A OrdabayevTimothy M Lohman
Dec 31, 2020·International Journal of Molecular Sciences·Aaron MorganChristopher Fischer
Dec 14, 2017·Biochemical and Biophysical Research Communications·Ryota HideseShinsuke Fujiwara
Jun 25, 2015·Microbiology Spectrum·J A Ruiz-MasóG Del Solar
Jun 4, 2021·Journal of Molecular Biology·Binh NguyenTimothy M Lohman

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