The informational content of DNA is held within the structures of the DNA bases. Modified DNA bases are considered “DNA damage” and have the potential to alter the coding and functional properties of DNA. DNA glycosylases play a key role in removing damaged DNA bases as an initiating step in base excision repair. My laboratory has focused extensively on the MutY/MUTYH glycosylase, which are an unusual [4Fe-4S] cluster containing enzyme that removes an undamaged adenine mispaired with damaged base 8-oxoguanine (OG). Our laboratory provided key information that led to the discovery of a new colorectal cancer predisposition mechanism, now referred to as MUTYH-associated polyposis (MAP). Notably, using modified substrates and modified enzymes in both in vitro and cellular assays we have revealed the key features of OG:A base pairs that allows MUTYH to distinguish them from other similar normal Watson-Crick bps. Structural studies of MutY bound to azaribose transition state (TS) analog containing DNA duplexes, along with determination of reaction stereochemistry, have rshown that MutY enzymes are “retaining” O-glycosidases. This new-for-MutY yet familiar mechanism may also be operative in related base excision repair glycosylases and provides a critical framework for analysis of MUTYH variants associated with MAP.<br/>Our laboratory has also focused on understanding the repair of the hydantoin DNA lesions identified and studied in detail by the James Flack Norris awardee Dr. Cynthia Burrows. The hydantoin lesions arise under conditions of high oxidative stress, and are more potently mutagenic than OG. Our laboratory has found that these lesions are removed by a variety of DNA glycosylases, and are the best-documented substrates for the human NEIL1 glycosylase. NEIL1 is an unusual glycosylase that works in a variety of DNA contexts, including single-stranded DNA and G-quadruplexes. In addition, we uncovered that there are two forms of the NEIL1 glycosylase resulting from mRNA editing (recoding) that have distinct differences in enzymatic processing of DNA base lesions. Our present work is focused on dissecting the molecular features of the repair of myriad of substrates, including the hydantoins, processed by NEIL1 and the consequences of NEIL1 recoding. This work illustrates the “fine-tuning” provided by DNA repair glycosylases to detect, mediate and respond to DNA base modifications.

DNA repair, organic chemistry & cancer: It’s all about the base

255th National Meeting (2018)

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