title: Insights into the substrate specificity of the MutT pyrophosphohydrolase using structural analogues of 8-oxo-2'-deoxyguanosine nucleotide.
abstract: The bacterial repair enzyme MutT hydrolyzes the damaged nucleotide OdGTP (the 5'-triphosphate derivative of 8-oxo-2'-deoxyguanosine; OdG), which is a known mutagen and has been linked to antibacterial action. Previous work has indicated important roles for the C8-oxygen, N7-hydrogen, and C2-exocyclic amine during OdGTP recognition by MutT. In order to gain a more nuanced understanding of the contribution of these three sites to the overall activity of MutT, we determined the reaction parameters for dGTP, OdGTP, and nine of their analogues using steady state kinetics. Our results indicate that overall high reaction efficiencies can be achieved despite altering any one of these sites. However, altering two or more sites leads to a significant decrease in efficiency. The data also suggest that, similar to another bacterial OdG repair enzyme, MutM, a specific carbonyl in the enzyme can not only promote activity by forming an active site hydrogen bond with the N7-hydrogen of OdGTP, but can also hinder activity through electrostatic repulsion with the N7-lone pair of dGTP.
authors.affiliation: Department of Chemistry, University of Richmond, Richmond, VA 23173, United States. Electronic address: mhamm@richmond.edu.
authors.forename: Michelle L
authors.initials: ML
authors.lastname: Hamm
authors.affiliation: Department of Chemistry, University of Richmond, Richmond, VA 23173, United States.
authors.forename: Emily J
authors.initials: EJ
authors.lastname: McFadden
authors.affiliation: Department of Chemistry, University of Richmond, Richmond, VA 23173, United States.
authors.forename: Michael
authors.initials: M
authors.lastname: Ghio
authors.affiliation: Department of Chemistry, University of Richmond, Richmond, VA 23173, United States.
authors.forename: Maria A M
authors.initials: MA
authors.lastname: Lindell
authors.affiliation: School of Chemistry and Materials Science, Rochester Institute of Technology, Rochester, NY 14623, United States.
authors.forename: Kenneth S
authors.initials: KS
authors.lastname: Gerien
authors.affiliation: School of Chemistry and Materials Science, Rochester Institute of Technology, Rochester, NY 14623, United States.
authors.forename: Suzanne F
authors.initials: SF
authors.lastname: O'Handley
mesh_terms: D000080242:8-Hydroxy-2'-Deoxyguanosine; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism*; D008958:Models, Molecular; D008968:Molecular Conformation; D011755:Pyrophosphatases / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D029968:Escherichia coli Proteins; D000080242:8-Hydroxy-2'-Deoxyguanosine; D011755:Pyrophosphatases; C052211:mutT protein, E coli; D003849:Deoxyguanosine
keywords: 8-Oxo-2′-deoxyguanosine; DNA repair; MutT; Oxidized nucleotides