Copper-dependent inhibition and oxidative inactivation with affinity cleavage of yeast glutathione reductase. 2014

Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
Department of Biochemistry, Aichi Medical University School of Medicine, Yazako-Karimata 1-1, Nagakute, Aichi, 480-1195, Japan.

Effects of copper on the activity and oxidative inactivation of yeast glutathione reductase were analyzed. Glutathione reductase from yeast was inhibited by cupric ion and more potently by cuprous ion. Copper ion inhibited the enzyme noncompetitively with respect to the substrate GSSG and NADPH. The Ki values of the enzyme for Cu(2+) and Cu(+) ion were determined to be 1 and 0.35 μM, respectively. Copper-dependent inactivation of glutathione reductase was also analyzed. Hydrogen peroxide and copper/ascorbate also caused an inactivation with the cleavage of peptide bond of the enzyme. The inactivation/fragmentation of the enzyme was prevented by addition of catalase, suggesting that hydroxyl radical produced through the cuprous ion-dependent reduction of oxygen is responsible for the inactivation/fragmentation of the enzyme. SDS-PAGE and TOF-MS analysis confirmed eight fragments, which were further determined to result from the cleavage of the Met17-Ser18, Asn20-Thr21, Glu251-Gly252, Ser420-Pro421, Pro421-Thr422 bonds of the enzyme by amino-terminal sequencing analysis. Based on the kinetic analysis and no protective effect of the substrates, GSSG and NADPH on the copper-mediated inactivation/fragmentation of the enzyme, copper binds to the sites apart from the substrate-sites, causing the peptide cleavage by hydroxyl radical. Copper-dependent oxidative inactivation/fragmentation of glutathione reductase can explain the prooxidant properties of copper under the in vivo conditions.

UI MeSH Term Description Entries
D007700 Kinetics The rate dynamics in chemical or physical systems.
D008969 Molecular Sequence Data Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories. Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D010084 Oxidation-Reduction A chemical reaction in which an electron is transferred from one molecule to another. The electron-donating molecule is the reducing agent or reductant; the electron-accepting molecule is the oxidizing agent or oxidant. Reducing and oxidizing agents function as conjugate reductant-oxidant pairs or redox pairs (Lehninger, Principles of Biochemistry, 1982, p471). Redox,Oxidation Reduction
D010446 Peptide Fragments Partial proteins formed by partial hydrolysis of complete proteins or generated through PROTEIN ENGINEERING techniques. Peptide Fragment,Fragment, Peptide,Fragments, Peptide
D005980 Glutathione Reductase Catalyzes the oxidation of GLUTATHIONE to GLUTATHIONE DISULFIDE in the presence of NADP+. Deficiency in the enzyme is associated with HEMOLYTIC ANEMIA. Formerly listed as EC 1.6.4.2. Glutathione-Disulfide Reductase,Reductase, Glutathione,Reductase, Glutathione-Disulfide
D000595 Amino Acid Sequence The order of amino acids as they occur in a polypeptide chain. This is referred to as the primary structure of proteins. It is of fundamental importance in determining PROTEIN CONFORMATION. Protein Structure, Primary,Amino Acid Sequences,Sequence, Amino Acid,Sequences, Amino Acid,Primary Protein Structure,Primary Protein Structures,Protein Structures, Primary,Structure, Primary Protein,Structures, Primary Protein
D001205 Ascorbic Acid A six carbon compound related to glucose. It is found naturally in citrus fruits and many vegetables. Ascorbic acid is an essential nutrient in human diets, and necessary to maintain connective tissue and bone. Its biologically active form, vitamin C, functions as a reducing agent and coenzyme in several metabolic pathways. Vitamin C is considered an antioxidant. Vitamin C,Ascorbic Acid, Monosodium Salt,Ferrous Ascorbate,Hybrin,L-Ascorbic Acid,Magnesium Ascorbate,Magnesium Ascorbicum,Magnesium di-L-Ascorbate,Magnorbin,Sodium Ascorbate,Acid, Ascorbic,Acid, L-Ascorbic,Ascorbate, Ferrous,Ascorbate, Magnesium,Ascorbate, Sodium,L Ascorbic Acid,Magnesium di L Ascorbate,di-L-Ascorbate, Magnesium
D012441 Saccharomyces cerevisiae A species of the genus SACCHAROMYCES, family Saccharomycetaceae, order Saccharomycetales, known as "baker's" or "brewer's" yeast. The dried form is used as a dietary supplement. Baker's Yeast,Brewer's Yeast,Candida robusta,S. cerevisiae,Saccharomyces capensis,Saccharomyces italicus,Saccharomyces oviformis,Saccharomyces uvarum var. melibiosus,Yeast, Baker's,Yeast, Brewer's,Baker Yeast,S cerevisiae,Baker's Yeasts,Yeast, Baker
D017665 Hydroxyl Radical The univalent radical OH. Hydroxyl radical is a potent oxidizing agent.
D059748 Proteolysis Cleavage of proteins into smaller peptides or amino acids either by PROTEASES or non-enzymatically (e.g., Hydrolysis). It does not include Protein Processing, Post-Translational. Protein Degradation,Protein Digestion,Degradation, Protein,Degradations, Protein,Digestion, Protein,Digestions, Protein,Protein Degradations,Protein Digestions,Proteolyses

Related Publications

Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
October 1996, Journal of enzyme inhibition,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
May 1996, Biochemical Society transactions,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
August 1995, European journal of biochemistry,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
November 2008, Comparative biochemistry and physiology. Part A, Molecular & integrative physiology,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
April 2021, Bulletin of environmental contamination and toxicology,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
January 1997, Chemical research in toxicology,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
March 2012, The Biochemical journal,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
October 1981, Indian journal of biochemistry & biophysics,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
May 1981, Biochimica et biophysica acta,
Keiko Murakami, and Ryoko Tsubouchi, and Minoru Fukayama, and Masataka Yoshino
May 2019, International journal of biological macromolecules,
Copied contents to your clipboard!