Kinetic Methods for Studying DNA Glycosylases Functioning in Base Excision Repair.

Abstract

Base excision repair (BER) is a conserved and ubiquitous pathway that is initiated by DNA glycosylases, which recognize and remove damaged or mismatched nucleobases, setting the stage for restoration of the correct DNA sequence by follow-on BER enzymes. DNA glycosylases employ a nucleotide-flipping step prior to cleavage of the N-glycosyl bond, and most exhibit slow release of the abasic DNA product and/or strong product inhibition. As such, studying the catalytic mechanism of these enzymes requires care in the design, execution, and interpretation of single- and multiple-turnover kinetics experiments, which is the topic of this chapter.