title: Movement of the β-hairpin in the third zinc-binding module of UvrA is required for DNA damage recognition.
abstract: Nucleotide excision repair (NER) is distinguished from other DNA repair pathways by its ability to process various DNA lesions. In bacterial NER, UvrA is the key protein that detects damage and initiates the downstream NER cascade. Although it is known that UvrA preferentially binds to damaged DNA, the mechanism for damage recognition is unclear. A β-hairpin in the third Zn-binding module (Zn3hp) of UvrA has been suggested to undergo a conformational change upon DNA binding, and proposed to be important for damage sensing. Here, we investigate the contribution of the dynamics in the Zn3hp structural element to various activities of UvrA during the early steps of NER. By restricting the movement of the Zn3hp using disulfide crosslinking, we showed that the movement of the Zn3hp is required for damage-specific binding, UvrB loading and ATPase activities of UvrA. We individually inactivated each of the nucleotide binding sites in UvrA to investigate its role in the movement of the Zn3hp. Our results suggest that the conformational change of the Zn3hp is controlled by ATP hydrolysis at the distal nucleotide binding site. We propose a bi-phasic damage inspection model of UvrA in which movement of the Zn3hp plays a key role in damage recognition.
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok 10400, Thailand; Center for Excellence in Protein and Enzyme Technology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
authors.forename: Thanyalak
authors.initials: T
authors.lastname: Kraithong
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok 10400, Thailand; Center for Excellence in Protein and Enzyme Technology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
authors.forename: Ketsaraphorn
authors.initials: K
authors.lastname: Channgam
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok 10400, Thailand; Center for Excellence for Shrimp Molecular Biology and Biotechnology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
authors.forename: Ornchuma
authors.initials: O
authors.lastname: Itsathitphaisarn
authors.affiliation: Department of Mathematics, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
authors.forename: Montip
authors.initials: M
authors.lastname: Tiensuwan
authors.affiliation: Department of Chemistry and Biochemistry, City College of New York, New York, NY 10031, USA; Ph.D. Programs in Biochemistry, Biology, and Chemistry, The Graduate Center of the City University of New York, New York, NY 10016, USA.
authors.forename: David
authors.initials: D
authors.lastname: Jeruzalmi
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok 10400, Thailand; Center for Excellence in Protein and Enzyme Technology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand. Electronic address: danaya.pak@mahidol.ac.th.
authors.forename: Danaya
authors.initials: D
authors.lastname: Pakotiprapha
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D006868:Hydrolysis; D009068:Movement; D017434:Protein Structure, Tertiary; D016335:Zinc Fingers
chemical_list: D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D000255:Adenosine Triphosphate; C068631:UvrA protein, E coli; D000251:Adenosine Triphosphatases
keywords: Disulfide crosslinking; NER; Nucleotide excision repair; UvrA