Characterization of the Redox Transition of the XRCC1 N-terminal Domain

• Published in: Structure (London) 2014-12, Vol.22 (12), p.1754-1763
• Main Authors: Gabel, Scott A., Smith, Cassandra E., Cuneo, Matthew J., Mueller, Geoffrey A., Kirby, Thomas W., DeRose, Eugene F., Krahn, Juno M., London, Robert E.
• Format: Article
• Language: English
• Subjects: Cell Line; DNA Repair; DNA-Binding Proteins - metabolism; Escherichia coli; Models, Molecular; Oxidation-Reduction; Protein Binding; Protein Folding; X-ray Repair Cross Complementing Protein 1
• ISSN: 0969-2126; 1878-4186
• DOI: 10.1016/j.str.2014.09.012

XRCC1, a scaffold protein involved in DNA repair, contains an N-terminal domain (X1NTD) that interacts specifically with DNA polymerase β. It was recently discovered that X1NTD contains a disulfide switch that allows it to adopt either of two metamorphic structures. In the present study, we demonstrate that formation of an N-terminal proline carbimate adduct resulting from the nonenzymatic reaction of Pro2 with CO2 is essential for stabilizing the oxidized structure, X1NTDox. The kinetic response of X1NTDred to H2O2, monitored by NMR, was determined to be very slow, consistent with involvement of the buried, kinetically trapped Cys12 residue, but was significantly accelerated by addition of protein disulfide isomerase or by Cu2+. NMR analysis of a sample containing the pol β polymerase domain, and both the reduced and oxidized forms of X1NTD, indicates that the oxidized form binds to the enzyme 25-fold more tightly than the reduced form. [Display omitted] •Oxidized XRCC1 is stabilized by formation of a Pro2-carbimate adduct•The XRCC1 redox transition can be catalyzed by protein disulfide isomerase•The affinity ratio of the oxidized/reduced forms of XRCC1 N-terminal for pol β is 25 Gabel et al. show that XRCC1, a scaffold protein involved in DNA repair, contains an N-terminal disulfide switch that regulates binding affinity for DNA polymerase β. The oxidized structure is dynamic, but is significantly stabilized by formation of an N-terminal carbimate adduct with available CO2.