title: Contribution of human manganese superoxide dismutase tyrosine 34 to structure and catalysis.
abstract: Superoxide dismutase (SOD) enzymes are critical in controlling levels of reactive oxygen species (ROS) that are linked to aging, cancer, and neurodegenerative disease. Superoxide (O(2)(*-)) produced during respiration is removed by the product of the SOD2 gene, the homotetrameric manganese superoxide dismutase (MnSOD). Here, we examine the structural and catalytic roles of the highly conserved active-site residue Tyr34, based upon structure-function studies of MnSOD enzymes with mutations at this site. Substitution of Tyr34 with five different amino acids retained the active-site protein structure and assembly but caused a substantial decrease in the catalytic rate constant for the reduction of superoxide. The rate constant for formation of the product inhibition complex also decreases but to a much lesser extent, resulting in a net increase in the level of product inhibited form of the mutant enzymes. Comparisons of crystal structures and catalytic rates also suggest that one mutation, Y34V, interrupts the hydrogen-bonded network, which is associated with a rapid dissociation of the product-inhibited complex. Notably, with three of the Tyr34 mutants, we also observe an intermediate in catalysis, which has not been reported previously. Thus, these mutants establish a means of trapping a catalytic intermediate that promises to help elucidate the mechanism of catalysis.
authors.affiliation: Department of Molecular Biology, The Scripps Research Institute, La Jolla, California 92037, USA. jjperry@scripps.edu
authors.forename: J Jefferson P
authors.initials: JJ
authors.lastname: Perry
authors.forename: Amy S
authors.initials: AS
authors.lastname: Hearn
authors.forename: Diane E
authors.initials: DE
authors.lastname: Cabelli
authors.forename: Harry S
authors.initials: HS
authors.lastname: Nick
authors.forename: John A
authors.initials: JA
authors.lastname: Tainer
authors.forename: David N
authors.initials: DN
authors.lastname: Silverman
mesh_terms: D002384:Catalysis; D020134:Catalytic Domain / Q000235:genetics; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D004252:DNA Mutational Analysis; D006801:Humans; D007700:Kinetics; D008345:Manganese / Q000737:chemistry* / Q000378:metabolism; D010084:Oxidation-Reduction; D013329:Structure-Activity Relationship; D013482:Superoxide Dismutase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry* / Q000235:genetics; D000099156:Superoxide Dismutase 2
chemical_list: D008345:Manganese; D013482:Superoxide Dismutase; D013481:Superoxides; D014443:Tyrosine; D000099156:Superoxide Dismutase 2