Key points are not available for this paper at this time.
Glutaredoxin (GRx, thioltransferase) is implicated in cellular redox regulation, and it is known for specific and efficient catalysis of reduction of protein-S-S-glutathione-mixed disulfides (protein-SSG) because of its remarkably low thiol pKa (≈3.5) and its ability to stabilize a catalytic S-glutathionyl intermediate (GRx-SSG). These unique properties suggested that GRx might also react with glutathione-thiyl radicals (GS⋅) and stabilize a disulfide anion radical intermediate (GRx-SSG⨪), thereby facilitating the conversion of GS⋅ to GSSG or transfer of GS⋅ to form protein-SSG. We found that GRx catalyzes GSSG formation in the presence of GS-thiyl radical generating systems (Fe2+/ADP/H2O2 + GSH or horseradish peroxidase/H2O2 + GSH). Catalysis is dependent on O2 and results in concomitant superoxide formation, and it is distinguished from glutathione peroxidase-like activity. With the horseradish peroxidase system and 35SGSH, GRx enhanced the rate of GS-radiolabel incorporation into GAPDH. GRx also enhanced the rate ofS-glutathionylation of glyceraldehyde-3-phosphate dehydrogenase with GSSG or S-nitrosoglutathione, but these glutathionyl donors were much less efficient. Both actin and protein-tyrosine phosphatase-1B were superior substrates for GRx-facilitated S-glutathionylation with GS-radical. These studies characterize GRx as a versatile catalyst, facilitating GS-radical scavenging and S-glutathionylation of redox signal mediators, consistent with a critical role in cellular regulation. Glutaredoxin (GRx, thioltransferase) is implicated in cellular redox regulation, and it is known for specific and efficient catalysis of reduction of protein-S-S-glutathione-mixed disulfides (protein-SSG) because of its remarkably low thiol pKa (≈3.5) and its ability to stabilize a catalytic S-glutathionyl intermediate (GRx-SSG). These unique properties suggested that GRx might also react with glutathione-thiyl radicals (GS⋅) and stabilize a disulfide anion radical intermediate (GRx-SSG⨪), thereby facilitating the conversion of GS⋅ to GSSG or transfer of GS⋅ to form protein-SSG. We found that GRx catalyzes GSSG formation in the presence of GS-thiyl radical generating systems (Fe2+/ADP/H2O2 + GSH or horseradish peroxidase/H2O2 + GSH). Catalysis is dependent on O2 and results in concomitant superoxide formation, and it is distinguished from glutathione peroxidase-like activity. With the horseradish peroxidase system and 35SGSH, GRx enhanced the rate of GS-radiolabel incorporation into GAPDH. GRx also enhanced the rate ofS-glutathionylation of glyceraldehyde-3-phosphate dehydrogenase with GSSG or S-nitrosoglutathione, but these glutathionyl donors were much less efficient. Both actin and protein-tyrosine phosphatase-1B were superior substrates for GRx-facilitated S-glutathionylation with GS-radical. These studies characterize GRx as a versatile catalyst, facilitating GS-radical scavenging and S-glutathionylation of redox signal mediators, consistent with a critical role in cellular regulation. glutaredoxin (thioltransferase) S-carboxymethyl bovine serum albumin S-nitrosoglutathione cytochrome c glyceraldehyde 3-phosphate dehydrogenase glutathione (reduced form) glutathione disulfide (oxidized form) horseradish peroxidase β-nicotinamide adenine dinucleotide phosphate protein-S-S-glutathione mixed disulfides protein tyrosine phosphatase 1B Human glutaredoxin (thioltransferase) (GRx,1 EC 1.8.4.2) is known for its unique properties of specific and efficient catalysis of deglutathionylation of protein-S-S-glutathione-mixed disulfides (protein-SSG) (1Gravina S.A. Mieyal J.J. Biochemistry. 1993; 32: 3368-3376Crossref PubMed Scopus (279) Google Scholar, 2Mieyal J.J. Gravina S.A. Mieyal P.A. Srinivasan U. Starke D.W. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 305-372Google Scholar, 3Yang Y. Jao S.-C. Nanduri S. Starke D.W. Mieyal J.J. Qin J. Biochemistry. 1998; 37: 17145-17156Crossref PubMed Scopus (134) Google Scholar, 4Chrestensen C.A. Starke D.W. Mieyal J.J. J. Biol. Chem. 2000; 275: 26556-26565Abstract Full Text Full Text PDF PubMed Scopus (279) Google Scholar). These catalytic properties of glutaredoxin have identified the enzyme for prominent roles in homeostasis of protein sulfhydryl groups both in a protective mode under overt oxidative stress associated with aging and various disease states including cardiovascular and neurodegenerative diseases, diabetes, AIDS, and cancer (2Mieyal J.J. Gravina S.A. Mieyal P.A. Srinivasan U. Starke D.W. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 305-372Google Scholar, 5Thomas J.A. Poland B. Honzatco R. Arch. Biochem. Biophys. 1995; 319: 1-9Crossref PubMed Scopus (367) Google Scholar, 6Cotgreave I.A. Gerdes R.C. Biochem. Biophys. Res. Commun. 1998; 242: 1-9Crossref PubMed Scopus (435) Google Scholar) and in a regulatory mode whereby reversible glutathionylation represents a mechanism of redox-activated signal transduction (7Sies H. Brigelius R. Graf P. Adv. Enzyme Regul. 1987; 26: 175-189Crossref PubMed Scopus (44) Google Scholar, 8Bandyopadhyay S. Starke D.W. Mieyal J.J. Gronostajski R.M. J. Biol. Chem. 1998; 273: 392-397Abstract Full Text Full Text PDF PubMed Scopus (129) Google Scholar, 9Barrett W.C. DeGnore J.P. Konig S. Fales H.M. Keng Y.F. Zhang Z.Y. Yim M.B. Chock P.B. Biochemistry. 1999; 38: 6699-6705Crossref PubMed Scopus (433) Google Scholar, 10Klatt P. Lamas S. Eur. J. Biochem. 2000; 267: 4928-4944Crossref PubMed Scopus (667) Google Scholar, 11Wang J. Boja E.S. Tan W. Tekel E. Fales H. English S.M. Mieyal J.J. Chock P.B. J. Biol. Chem. 2001; 276: 47763-47766Abstract Full Text Full Text PDF PubMed Scopus (291) Google Scholar). These physiological roles are supported further by the documentation that glutaredoxin accounts for essentially all of cellular protein-SSG deglutathionylase activity in mammalian cells (4Chrestensen C.A. Starke D.W. Mieyal J.J. J. Biol. Chem. 2000; 275: 26556-26565Abstract Full Text Full Text PDF PubMed Scopus (279) Google Scholar, 12Gravina, S. A., Characterization and Kinetic Mechanism of Thioltransferase.Ph.D. thesis, 1993, Department of Pharmacology, Case Western Reserve University.Google Scholar, 13Jung C.H. Thomas J.A. Arch. Biochem. Biophys. 1996; 335: 61-72Crossref PubMed Scopus (164) Google Scholar) and its inactivation by cadmium is correlated with inhibition of intracellular deglutathionylase activity (4Chrestensen C.A. Starke D.W. Mieyal J.J. J. Biol. Chem. 2000; 275: 26556-26565Abstract Full Text Full Text PDF PubMed Scopus (279) Google Scholar, 11Wang J. Boja E.S. Tan W. Tekel E. Fales H. English S.M. Mieyal J.J. Chock P.B. J. Biol. Chem. 2001; 276: 47763-47766Abstract Full Text Full Text PDF PubMed Scopus (291) Google Scholar). Although reversible formation of protein-SSG is a prevalent form of protein sulfhydryl modification, mechanisms of protein-SSG formation are not resolved. Unless intracellular GSSG concentrations reach unusually high levels, GSSG is unlikely to be the mediator of protein-SSG formation based on typical redox potentials for cysteine residues (14Ziegler D.M. Annu. Rev. Biochem. 1985; 54: 305-329Crossref PubMed Scopus (726) Google Scholar, 15Gilbert H.F. Methods Enzymol. 1995; 251: 8-28Crossref PubMed Scopus (504) Google Scholar). Consequently, glutathione-thiyl radical andS-nitrosoglutathione (GS-NO) have been considered as potential alternative mediators (5Thomas J.A. Poland B. Honzatco R. Arch. Biochem. Biophys. 1995; 319: 1-9Crossref PubMed Scopus (367) Google Scholar, 6Cotgreave I.A. Gerdes R.C. Biochem. Biophys. Res. Commun. 1998; 242: 1-9Crossref PubMed Scopus (435) Google Scholar, 10Klatt P. Lamas S. Eur. J. Biochem. 2000; 267: 4928-4944Crossref PubMed Scopus (667) Google Scholar). In this context, we considered the unusually low pKa (=3.5) of the active site cysteine of glutaredoxin (16Mieyal J.J. Starke D.W. Gravina S.A. Hocevar B. Biochemistry. 1991; 30: 8883-8891Crossref PubMed Scopus (89) Google Scholar, 17Srinivasan U. Mieyal P.A. Mieyal J.J. Biochemistry. 1997; 36: 3199-3206Crossref PubMed Scopus (109) Google Scholar) and the selective stabilization of the glutathionyl moiety in the glutaredoxin-SSG catalytic intermediate (1Gravina S.A. Mieyal J.J. Biochemistry. 1993; 32: 3368-3376Crossref PubMed Scopus (279) Google Scholar, 2Mieyal J.J. Gravina S.A. Mieyal P.A. Srinivasan U. Starke D.W. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 305-372Google Scholar, 3Yang Y. Jao S.-C. Nanduri S. Starke D.W. Mieyal J.J. Qin J. Biochemistry. 1998; 37: 17145-17156Crossref PubMed Scopus (134) Google Scholar) along with the known ability of thiyl radicals to be stabilized by the formation of disulfide anion radicals (18Wardman P. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 1-19Google Scholar, 19Schoneich C. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 21-47Google Scholar). Accordingly, we reasoned that glutaredoxin might react preferentially with the glutathione-thiyl radical to form a glutaredoxin-S-S-glutathione-disulfide anion radical (GRx-SSG⨪) and that this enzyme intermediate could facilitate transfer of the GS-radical either to form GSSG or protein-SSG adducts. To test this hypothesis, two GS-radical generating systems were used,i.e. Fe(II)-ADP/H2O2 + GSH (20Floyd R.A. Lewis C.A. Biochemistry. 1983; 22: 2645-2649Crossref PubMed Scopus (154) Google Scholar, 21Floyd R.A. Arch. Biochem. Biophys. 1983; 225: 263-270Crossref PubMed Scopus (112) Google Scholar) and HRP/H2O2 + GSH (22Harman L.S. Carver D.K. Schreiber J. Mason R.P. J. Biol. Chem. 1986; 261: 1642-1648Abstract Full Text PDF PubMed Google Scholar). Here we report that glutaredoxin catalyzes the formation of GSSG in the GS-radical generating systems. The catalysis of GSSG formation is dependent on molecular oxygen, and it is distinguished from glutathione peroxidase-like activity. Moreover, glutaredoxin enhanced the rate ofS-glutathionylation of GAPDH in the presence of GS-radicals. This model reaction mimics the intracellular formation of GAPDH-SSG under oxidative conditions where GSSG content is not substantially changed (23Chai Y.C. Hendrich S. Thomas J.A. Arch. Biochem. Biophys. 1994; 310: 264-272Crossref PubMed Scopus (76) Google Scholar, 24Chai Y.C. Ashraf S.S. Rokutan K. Johnston Jr., R.B. Thomas J.A. Arch. Biochem. Biophys. 1994; 310: 273-281Crossref PubMed Scopus (199) Google Scholar, 25Ravichandran V. Seres T. Moriguchi T. Thomas J.A. Johnston Jr., R.B. J. Biol. Chem. 1994; 269: 25010-25015Abstract Full Text PDF PubMed Google Scholar). Glutaredoxin also enhanced the rate of formation of GAPDH-SSG when GSSG or GS-NO were used as the glutathionyl donors, but these reactions were much less efficient than the GS-radical transfer reaction. In comparison to GAPDH, both actin and PTP1B were found to be superior substrates for GRx-facilitated S-glutathionylation under GS-radical transfer conditions. Both actin and PTP1B are implicated in redox signal transduction, and both have been shown to be susceptible to intracellular regulation via reversible glutathionylation, probably involving glutaredoxin as the deglutathionylation catalyst (9Barrett W.C. DeGnore J.P. Konig S. Fales H.M. Keng Y.F. Zhang Z.Y. Yim M.B. Chock P.B. Biochemistry. 1999; 38: 6699-6705Crossref PubMed Scopus (433) Google Scholar, 11Wang J. Boja E.S. Tan W. Tekel E. Fales H. English S.M. Mieyal J.J. Chock P.B. J. Biol. Chem. 2001; 276: 47763-47766Abstract Full Text Full Text PDF PubMed Scopus (291) Google Scholar). Our current findings further characterize glutaredoxin as a versatile catalyst capable of facilitating S-glutathionylation of redox signal mediators. Water was initially purified by reverse osmosis. It was further purified by a Millipore MilliQ system. it was with of to and phosphate were from glutathione from glutathione peroxidase from bovine and were from GAPDH, horseradish cytochrome and were from was from Glutaredoxin was and purified as Y. Jao S.-C. Nanduri S. Starke D.W. Mieyal J.J. Qin J. Biochemistry. 1998; 37: 17145-17156Crossref PubMed Scopus (134) Google Scholar). phosphate was used in all of the from that was by phosphate to phosphate the was of were by of and the reaction phosphate various of GSSG and and were with the and glutaredoxin and were in to GSSG was the for in the were and used to as GSSG Glutaredoxin enzyme was as in the and of formation and on enzyme were were as with the that all of the were for with In and were to results with in the or presence of the system were were as with the that cytochrome c was to the of GSSG formation were as in were in a and of cytochrome c were radicals were by horseradish peroxidase and GSH from the studies of (22Harman L.S. Carver D.K. Schreiber J. Mason R.P. J. Biol. Chem. 1986; 261: 1642-1648Abstract Full Text PDF PubMed Google Scholar) and Mason R.P. Methods Enzymol. Scholar). The the incorporation of into GAPDH. GAPDH, Glutaredoxin was in various concentrations as and was to the reaction. were either or various reaction by to the The were with and with and into and were as a protein to facilitate of GAPDH. The of GAPDH-SSG disulfide was to the specific from GSH content and of in reaction was to GAPDH-SSG formation by of with disulfide was by GSSG with a of was as R.M. Biochem. PubMed Scopus Google Scholar) by with of and for We in model with GSH that this reaction conversion based on the of GS-NO Biochemistry. 1999; Google Scholar). and GSSG were as for ability to S-glutathionylation of GAPDH in the or presence of and GAPDH, and of the was to the reaction as or or or or GS-thiyl radical as The of probably represents based on the of GSSG in in the GS-radical generating system. The reactions were with and was to the specific of the glutathionyl as of in cells is implicated in signal transduction mechanisms as as of the mediators of these is and under active To the of glutathione thiyl radical and the potential roles of glutaredoxin in these we used the known system of the and (20Floyd R.A. Lewis C.A. Biochemistry. 1983; 22: 2645-2649Crossref PubMed Scopus (154) Google Scholar, 21Floyd R.A. Arch. Biochem. Biophys. 1983; 225: 263-270Crossref PubMed Scopus (112) Google Scholar, D.W. Y. Mieyal J.J. Biol. 1997; PubMed Scopus Google Scholar) to radicals in the or presence of GSH and radical in the of GSH was by the conversion of to as D.K. Biochem. Biophys. Res. Commun. PubMed Scopus Google and the radical transfer reaction of radicals with GSH to glutathionyl thiyl radicals (18Wardman P. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 1-19Google Scholar) was by GSH inhibition of formation in the Fe(II)-ADP/H2O2 system with not Although glutathionyl radical form the disulfide anion radical and react with to form superoxide and GSSG M.B. Chock P.B. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar, Methods Enzymol. 1995; 251: PubMed Scopus Google this reaction is P. C. Methods Enzymol. 1995; 251: PubMed Scopus Google Scholar) the of the formation of GSSG is a that is for consistent with a catalytic role for the Accordingly, the reaction is enzyme dependent and it on the enzyme for enzyme is not of on glutaredoxin Glutaredoxin was with and GSSG in phosphate were by of a and and were and GSSG was from the of the to the of comparison of glutathione peroxidase and glutaredoxin as of GSSG formation from GSH and in the of GS-radicals. were as with the that the was GS-radical formation and the was as glutathione peroxidase or glutaredoxin was as potential The formation of GSSG is distinguished from a glutathione peroxidase-like activity of because glutaredoxin on the rate of formation of GSSG with and GSH in the of the In glutathione peroxidase under the conditions in the of GSSG formation as To the reaction the of molecular oxygen, we the of that the reaction O2 for efficient and formation of superoxide mechanism of GS-radical scavenging by The the mechanism of glutaredoxin catalysis of GS-radical To this we concomitant reduction of is for superoxide formation of the of GSSG formation and superoxide formation is by the presence of with reaction with superoxide is by the rate of c reduction that as the c was of oxidative stress in cells is the of a protective mechanism (2Mieyal J.J. Gravina S.A. Mieyal P.A. Srinivasan U. Starke D.W. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 305-372Google Scholar, 5Thomas J.A. Poland B. Honzatco R. Arch. Biochem. Biophys. 1995; 319: 1-9Crossref PubMed Scopus (367) Google of specific been also of in as and probably represents a mode of regulation (9Barrett W.C. DeGnore J.P. Konig S. Fales H.M. Keng Y.F. Zhang Z.Y. Yim M.B. Chock P.B. Biochemistry. 1999; 38: 6699-6705Crossref PubMed Scopus (433) Google Scholar, 11Wang J. Boja E.S. Tan W. Tekel E. Fales H. English S.M. Mieyal J.J. Chock P.B. J. Biol. Chem. 2001; 276: 47763-47766Abstract Full Text Full Text PDF PubMed Scopus (291) Google Scholar). Although with specific cysteine residues that are to reversible glutathionylation have been (2Mieyal J.J. Gravina S.A. Mieyal P.A. Srinivasan U. Starke D.W. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 305-372Google Scholar, 10Klatt P. Lamas S. Eur. J. Biochem. 2000; 267: 4928-4944Crossref PubMed Scopus (667) Google the of formation of protein-SSG is protein glutathionylation been in various is glyceraldehyde-3-phosphate This protein been to as GAPDH-SSG in cells oxidative the GSSG not substantially (23Chai Y.C. Hendrich S. Thomas J.A. Arch. Biochem. Biophys. 1994; 310: 264-272Crossref PubMed Scopus (76) Google Scholar, 24Chai Y.C. Ashraf S.S. Rokutan K. Johnston Jr., R.B. Thomas J.A. Arch. Biochem. Biophys. 1994; 310: 273-281Crossref PubMed Scopus (199) Google Scholar, 25Ravichandran V. Seres T. Moriguchi T. Thomas J.A. Johnston Jr., R.B. J. Biol. Chem. 1994; 269: 25010-25015Abstract Full Text PDF PubMed Google Scholar, P. T. I.A. Eur. J. Biochem. 1994; PubMed Scopus Google that a form of the glutathione moiety than GSSG was the mediator of we the of GS-radical to as the of protein-SSG formation and glutaredoxin facilitate the reaction. In this the GS-radical generating system was horseradish as and GSH as This system to reaction of with GAPDH C. Gerdes R. I.A. Biochem. Biophys. Res. Commun. 1998; PubMed Scopus Google because we could a of than that for the system and a of GS-radical the reaction of GAPDH could be under conditions to a cellular low this GS-radical generating glutaredoxin the rate of formation in a and of GAPDH-SSG formation when the glutaredoxin enzyme was it was or when either or was from the reaction not we that the of GAPDH-SSG formation glutaredoxin and it is dependent on GS-radical that of not GAPDH-SSG formation from in the presence of GRx in to the on GSSG formation in the of less GAPDH-SSG is under conditions. the of GAPDH glutathionylation by GRx is in the of This for the glutathionylation of mechanism of GS-radical transfer by The a for S-glutathionylation of by GS-radicals. alternative less is shown GSSG and the glutathione-thiyl been on GS-NO as a potential mediator of protein-SSG formation P. Lamas S. Eur. J. Biochem. 2000; 267: 4928-4944Crossref PubMed Scopus (667) Google Scholar, S. H. B. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). we the ability of and GS-NO to GAPDH-SSG formation in the and presence of glutaredoxin Although both GSSG and GS-NO could for the GS-radical generating system and glutaredoxin GAPDH-SSG formation, the of these reactions was the in the of the reactions for GS-NO and GSSG the of GS-radical less than of the of GAPDH-SSG in the GS-radical reaction. a high were GS-NO and GSSG as or donors for as GS-radical with various glutathionyl in and GSSG were for ability to glutathionylation of GAPDH in the or presence of and GAPDH, and the the was to the reaction. The of probably represents based on the of GSSG in in the GS-radical generating system. The reactions were with and was to the specific of the glutathionyl as under in a and GSSG were for ability to glutathionylation of GAPDH in the or presence of and GAPDH, and the the was to the reaction. The of probably represents based on the of GSSG in in the GS-radical generating system. The reactions were with and was to the specific of the glutathionyl as under in the intracellular S-glutathionylation of PTP1B and actin in to a physiological redox have been the glutathionylation mechanism to be resolved. we also the of glutaredoxin to formation of and to GAPDH-SSG Both actin and PTP1B were much by GS-radical transfer with GAPDH. of the the in in the protein-SSG formation, that actin and PTP1B are by the GRx and GS-radical system and than GAPDH. been M.B. Chock P.B. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar, Methods Enzymol. 1995; 251: PubMed Scopus Google Scholar, Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: Scholar) that the role of GSH in the of radicals The intermediate formation of the GS-thiyl radical and the the radical is by reaction with molecular to the superoxide anion is to and O2 by the of superoxide and glutathione peroxidase to the formation of the anion radical is because of a low rate and the that GSH is in the form physiological the reaction to formation of superoxide is by the rate for P. C. Methods Enzymol. 1995; 251: PubMed Scopus Google Scholar). In this we the that the in this reaction could be by reaction involving glutaredoxin The of of GSSG formation by GRx catalytic of the the of of GSSG formation rate on GRx a of a typical cellular of GRx D.W. Y. Mieyal J.J. Biol. 1997; PubMed Scopus Google be to GSSG in In of the GS-radical could much because its reaction with GRx much than the reaction. It that the properties of glutaredoxin that this catalytic activity for scavenging the GS-radical be to that are for its efficient catalysis of protein-SSG for the glutathionyl moiety (1Gravina S.A. Mieyal J.J. Biochemistry. 1993; 32: 3368-3376Crossref PubMed Scopus (279) Google Scholar, 3Yang Y. Jao S.-C. Nanduri S. Starke D.W. Mieyal J.J. Qin J. Biochemistry. 1998; 37: 17145-17156Crossref PubMed Scopus (134) Google and unusually low pKa of the active site cysteine thiol of (16Mieyal J.J. Starke D.W. Gravina S.A. Hocevar B. Biochemistry. 1991; 30: 8883-8891Crossref PubMed Scopus (89) Google Scholar, 17Srinivasan U. Mieyal P.A. Mieyal J.J. Biochemistry. 1997; 36: 3199-3206Crossref PubMed Scopus (109) Google Scholar). These properties stabilization of a disulfide anion radical intermediate as in To the disulfide anion radical intermediate might be by with a GS-radical or of molecular we the of The for GRx catalysis of GS-radical scavenging are consistent with the of is to the reaction we for GRx catalysis of reduction of disulfides (2Mieyal J.J. Gravina S.A. Mieyal P.A. Srinivasan U. Starke D.W. Packer L. Cadenas E. Biothiols in Health and Disease. Marcel Dekker, Inc., New York1995: 305-372Google Scholar). formation of GSSG is dependent on conditions that the glutathione-thiyl and catalysis is by the of molecular With O2 concomitant formation of superoxide along with the GSSG was shown by reduction of cytochrome than reduction of c and GSSG formation was probably because of and for reaction with the Accordingly, the of to of reduction a mechanism for scavenging radicals cells where GSH is in and as the the GS-radical. glutaredoxin the GS-radical as the stabilized disulfide anion radical intermediate that react with molecular oxygen, in conversion of the radical to superoxide anion The superoxide radical be by the of superoxide and glutathione peroxidase or the peroxidase system to molecular O2 and as the of the scavenging radical be by GSH to the catalytic of glutaredoxin and superoxide of on of was R. Biochemistry. 1995; PubMed Scopus Google Scholar). is a in reversible S-glutathionylation as a mechanism of signal transduction I.A. Gerdes R.C. Biochem. Biophys. Res. Commun. 1998; 242: 1-9Crossref PubMed Scopus (435) Google Scholar, H. Brigelius R. Graf P. Adv. Enzyme Regul. 1987; 26: 175-189Crossref PubMed Scopus (44) Google Scholar, 10Klatt P. Lamas S. Eur. J. Biochem. 2000; 267: 4928-4944Crossref PubMed Scopus (667) Google Scholar, J.J. Chock P.B. Chock P. in New Scholar). Although much this are the prominent is are mechanisms for formation of specific protein-SSG as This to this and for the presence of intracellular protein-SSG under conditions where GSSG not to consistent with glutathione-thiyl radical been suggested as alternative (5Thomas J.A. Poland B. Honzatco R. Arch. Biochem. Biophys. 1995; 319: 1-9Crossref PubMed Scopus (367) Google Scholar, 10Klatt P. Lamas S. Eur. J. Biochem. 2000; 267: 4928-4944Crossref PubMed Scopus (667) Google Scholar) consistent with this hypothesis, intracellular GS-thiyl radicals have been and identified by H. Yim H. Chock P.B. Yim M.B. U. S. 1995; PubMed Scopus Google Scholar, S. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus (44) Google Scholar). In this GAPDH was used as the model to glutaredoxin catalyzes protein-SSG formation, and GS-NO and GSSG were as alternative shown in formation of GAPDH-SSG was efficient when GS-radical was the of the GS-radical GSSG Although the mechanisms by GRx GAPDH-SSG formation with of these donors are the comparison donors that a reaction involving radical is and that of GS-radical the is efficient than of GS-NO either in the or presence of on current is a model for GRx facilitate protein-SSG studies are to the the conditions used for the the formation of GS-radical probably is rate for the reaction. Accordingly, glutaredoxin react with the to form the disulfide anion radical intermediate This intermediate could a from the to form thiyl radical and GSH of the reaction a radical reaction the protein-SSG and the glutaredoxin The reaction two for protein-SSG the might react with the intermediate to form and transfer the radical to molecular to protein-SSG and superoxide This alternative not be the protein properties to glutaredoxin pKa for the moiety and stabilization site for the glutathionyl GAPDH as a the rate of formation of protein-SSG was enhanced under conditions of This is consistent with a of the intermediate for reaction with the moiety when the reaction with O2 is of protein-SSG also be by GRx catalysis of To this with the GAPDH reaction GAPDH were and as substrates in a typical deglutathionylation for GRx in the of the GS-radical generating system. These were of from in a to the GRx not as the of protein-SSG in the reaction its rate of deglutathionylation a was In this we also the of actin and PTP1B as of S-glutathionylation by the GS-radical system because both of these have been to have intracellular by S-glutathionylation (9Barrett W.C. DeGnore J.P. Konig S. Fales H.M. Keng Y.F. Zhang Z.Y. Yim M.B. Chock P.B. Biochemistry. 1999; 38: 6699-6705Crossref PubMed Scopus (433) Google Scholar, 11Wang J. Boja E.S. Tan W. Tekel E. Fales H. English S.M. Mieyal J.J. Chock P.B. J. Biol. Chem. 2001; 276: 47763-47766Abstract Full Text Full Text PDF PubMed Scopus (291) Google Scholar). of and this that the of these under the conditions of was efficient to the deglutathionylation reaction in the system as for the current intracellular conditions where a of a low of when a redox is PTP1B was by GRx Although the of PTP1B and GAPDH with the anion radical intermediate could be to low pKa the be to actin cysteine a pKa J. Boja E.S. Tan W. Tekel E. Fales H. English S.M. Mieyal J.J. Chock P.B. J. Biol. Chem. 2001; 276: 47763-47766Abstract Full Text Full Text PDF PubMed Scopus (291) Google Scholar). the of of the as substrates for the two reactions by It protein S-glutathionylation and it is in the that glutaredoxin with catalyzes the formation of specific protein-SSG via transfer of the glutathione-thiyl radical is supported by the current studies that of The catalytic properties of glutaredoxin by the current studies characterize it as a versatile enzyme for a of the that glutaredoxin a role in sulfhydryl homeostasis and redox signal We for a of actin and Zhang for We also for critical of the
Starke et al. (Tue,) studied this question.