Structural and biochemical study on the covalent bond formation mechanism of a novel uracil DNA glycosylase UDGX

Abstract

Uracil DNA glycosylases (UDGs) are important DNA repair enzymes that excise uracil from DNA, yielding an abasic site. Recently, UdgX, an unconventional UDG with extremely tight binding to uracil-containing DNA was discovered. The structure of UdgX from Mycobacterium smegmatis in complex with DNA shows an overall similarity to that of the family 4 UDGs except for a protruding loop at the entrance of the uracil binding pocket. Surprisingly, H109 in the loop was found to make a covalent bond to the abasic site to form a stable intermediate, while the excised uracil remained in the active-site pocket. H109 functions as a nucleophile to attack the oxocarbenium ion, substituting the catalytic water molecule found in other UDGs. This change from the catalytic water attack to a direct nucleophilic attack by the histidine residue is unprecedented. UdgX utilizes a unique mechanism of protecting cytotoxic abasic sites from exposure to the cellular environment.