The base excision repair glycosylase MUTYH and its homologues are responsible for the removal of adenine (A) when base paired opposite a common product of oxidative DNA damage, 8-oxo-7,8-dihydroguanine (OG).1 When MUTYH is unable to perform base excision, it can result in changes in genetic information and therefore mutagenic phenotypes.1 This mutagenesis has been observed within the excised adenomas and carcinomas of colorectal cancer patients in a disease, known as MUTYH-associated polyposis (MAP).1 MAP is characterized by the inheritance of misfunctional variant forms of MUTYH, resulting in an accumulation of G:C to T:A transversions that often occur at critical cell regulator genes,.1 Many of these cancer-associated variants have amino acids changes that occur at positions surrounding the enzyme’s metal cofactor binding sites.1 One of these cofactors found in all MUTYH homologues is a [4Fe-4S]2+ cluster coordinated to four Cys residues located in the N-terminal catalytic domain.2 Recently, our lab also discovered that there is a second metal binding site in mammalian MUTYH homologues: three additional highly conserved Cys residues that coordinate to Zn2+ in the interdomain connector (IDC) region.3 The IDC region serves as the docking site for many DNA and cell regulator proteins such as the Rad9-Rad1-Hus1 binding complex.3 Therefore it is pertinent to understand the function of these metal cofactors and how these metal binding sites influence overall DNA repair.3 We utilized bioinformatics and a massive sequence alignment to in search of a possible fourth ligand for the Zn2+ and to provide insight into its evolutional significance. Also, with the aid of computational modeling we have generated a homology based structure optimizing cofactor binding within MUTYH in order to view spatial possibilities for a fourth ligand.. Lastly, we also explored possible functionalities of the newly discovered Zn2+ binding site by assessing the enzyme’s affinity for Zn2+, probing its possible structural role via circular dichroism, and evaluating how oxidative stress can influence Zn2+ binding.

Determining the ligand coordination sphere and functionality of the Zn2+ linchpin motif of the DNA repair glycosylase MUTYH

251st National Meeting (2016)

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