Characterization of binding constants and repair quantum yields of cyclobutane pyrimidine dimer by various class II DNA photolyase mutations

Abstract

Photolyase is a flavoprotein that repairs DNA damage caused by solar ultraviolet radiation. The repair quantum yields of the class I photolyases are around 80%, and for the class II photolyases are around 50%. In this research, we designed six key residues at the active site of class II MmPL from Methanosarcina mazei to systematically characterize the binding constants and quantum yields. We used titration experiments to detect the binding affinity of MmPL wild-type and the mutants with the oligomer containing T<>T lesion. Based on the dissociation constants, we can calculate the enzyme-substrate complex concentrations and obtained the repair quantum yields. Through mutation, we unexpectedly obtained higher binding affinity and higher repair quantum yields than the wild type, suggesting the other role of some residues. These results are critical to studies of ultrafast repair dynamics to solve the repair photocycles.