Mechanochemical regulations of RPA's binding to ssDNA

Scientific Reports
Jin ChenJie Yan

Abstract

Replication protein A (RPA) is a ubiquitous eukaryotic single-stranded DNA (ssDNA) binding protein that serves to protect ssDNA from degradation and annealing, and as a template for recruitment of many downstream factors in virtually all DNA transactions in cell. During many of these transactions, DNA is tethered and is likely subject to force. Previous studies of RPA's binding behavior on ssDNA were conducted in the absence of force; therefore the RPA-ssDNA conformations regulated by force remain unclear. Here, using a combination of atomic force microscopy imaging and mechanical manipulation of single ssDNA tethers, we show that force mediates a switch of the RPA bound ssDNA from amorphous aggregation to a much more regular extended conformation. Further, we found an interesting non-monotonic dependence of the binding affinity on monovalent salt concentration in the presence of force. In addition, we discovered that zinc in micromolar concentrations drives ssDNA to a unique, highly stiff and more compact state. These results provide new mechanochemical insights into the influences and the mechanisms of action of RPA on large single ssDNA.

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Citations

Apr 6, 2017·PloS One·Javier JarilloFrancisco J Cao
Aug 3, 2017·Nanotechnology·Xiaodan ZhaoJie Yan
Aug 29, 2020·Critical Reviews in Biochemistry and Molecular Biology·Colleen C Caldwell, Maria Spies
Jul 28, 2019·The Journal of Biological Chemistry·Qing-Man WangXi-Miao Hou
Feb 24, 2019·Current Genetics·Shuqi LiQing Li
Nov 5, 2019·Current Opinion in Chemical Biology·Xiaodan ZhaoJie Yan
Jun 10, 2020·Mutation Research·Colleen J LawrimoreKerry Bloom
Mar 1, 2018·CAAI Transactions on Intelligence Technology·Zhongyuan HuangZhe Wang

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Methods Mentioned

BETA
X-ray
electron
atomic force microscopy
AFM
saturation binding
PCR

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