title: The effect of mixed-function oxidation of enzymes on their susceptibility to degradation by a nonlysosomal cysteine proteinase.
abstract: Mixed-function oxidation of Escherichia coli glutamine synthetase by ascorbate, oxygen, and iron has previously been shown to cause inactivation of the enzyme and enhanced susceptibility to proteolytic attack by a variety of proteases. One of these proteases, from rat liver, is a high molecular weight cysteine proteinase which does not degrade native glutamine synthetase at neutral pH. Although inactive, the oxidized glutamine synthetase preparations used in this study were only partially degraded by this proteinase. Some of the subunits were degraded to acid soluble products with no detectable intermediates; the remaining subunits had not become susceptible to proteolytic attack during the limited exposure to the ascorbate mixed-function oxidation system. Several mammalian enzymes which are known to be inactivated by mixed-function oxidation were tested as substrates for the proteinase. Native rabbit muscle enolase and pyruvate kinase were resistant to degradation, but their oxidatively inactivated forms were degraded. Oxidized phosphoglycerate kinase and creatine kinase were also preferentially degraded. Moreover, trypsin degraded oxidized preparations of all of these enzymes faster than control preparations. Oxidative inactivation of superoxide dismutase by hydrogen peroxide caused a slight increase in susceptibility to proteolytic attack, but the enzyme was still relatively resistant to degradation both by the cysteine proteinase and by trypsin. Although oxidation conditions may not have been optimal for demonstrating enhanced proteolytic susceptibility, the results do indicate that mixed-function oxidation can render some mammalian enzymes, as well as bacterial glutamine synthetase, susceptible to degradation. Mixed-function oxidation of these proteins may be a mechanism of marking them for intracellular turnover.
authors.forename: A J
authors.initials: AJ
authors.lastname: Rivett
mesh_terms: D000818:Animals; D002417:Cattle; D003546:Cysteine Endopeptidases; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism*; D004798:Enzymes / Q000378:metabolism*; D005974:Glutamate-Ammonia Ligase / Q000378:metabolism; D006868:Hydrolysis; D008099:Liver / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D008970:Molecular Weight; D010084:Oxidation-Reduction; D011817:Rabbits; D014357:Trypsin / Q000378:metabolism
chemical_list: D004798:Enzymes; D006899:Mixed Function Oxygenases; D010450:Endopeptidases; D014357:Trypsin; D003546:Cysteine Endopeptidases; D005974:Glutamate-Ammonia Ligase