Catalysis of Thiol/Disulfide Exchange

Abstract

Glutaredoxin (Grx) and protein-disulfide isomerase (PDI) are members of the thioredoxin superfamily of thiol/disulfide exchange catalysts. Thermodynamically, rat PDI is a 600-fold better oxidizing agent than Grx1 from Escherichia coli. Despite that, Grx1 is a surprisingly good protein oxidase. It catalyzes protein disulfide formation in a redox buffer with an initial velocity that is 30-fold faster than PDI. Catalysis of protein and peptide oxidation by the individual catalytic domains of PDI and by a Grx1-PDI chimera show that differences in active site chemistry are fundamental to their oxidase activity. Mutations in the active site cysteines reveal that Grx1 needs only one cysteine to catalyze rapid substrate oxidation, whereas PDI requires both cysteines. Grx1 is a good oxidase because of the high reactivity of a Grx1-glutathione mixed disulfide, and PDI is a good oxidase because of the high reactivity of the disulfide between the two active site cysteines. As a protein disulfide reductase, Grx1 is also superior to PDI. It catalyzes the reduction of nonnative disulfides in scrambled ribonuclease and protein-glutathione mixed disulfides 30–180 times faster than PDI. A multidomain structure is necessary for PDI to catalyze effective protein reduction; however, placing Grx1 into the PDI multidomain structure does not enhance its already high reductase activity. Grx1 and PDI have both found mechanisms to enhance active site reactivity toward proteins, particularly in the kinetically difficult direction: Grx1 by providing a reactive glutathione mixed disulfide to supplement its oxidase activity and PDI by utilizing its multidomain structure to supplement its reductase activity. Glutaredoxin (Grx) and protein-disulfide isomerase (PDI) are members of the thioredoxin superfamily of thiol/disulfide exchange catalysts. Thermodynamically, rat PDI is a 600-fold better oxidizing agent than Grx1 from Escherichia coli. Despite that, Grx1 is a surprisingly good protein oxidase. It catalyzes protein disulfide formation in a redox buffer with an initial velocity that is 30-fold faster than PDI. Catalysis of protein and peptide oxidation by the individual catalytic domains of PDI and by a Grx1-PDI chimera show that differences in active site chemistry are fundamental to their oxidase activity. Mutations in the active site cysteines reveal that Grx1 needs only one cysteine to catalyze rapid substrate oxidation, whereas PDI requires both cysteines. Grx1 is a good oxidase because of the high reactivity of a Grx1-glutathione mixed disulfide, and PDI is a good oxidase because of the high reactivity of the disulfide between the two active site cysteines. As a protein disulfide reductase, Grx1 is also superior to PDI. It catalyzes the reduction of nonnative disulfides in scrambled ribonuclease and protein-glutathione mixed disulfides 30–180 times faster than PDI. A multidomain structure is necessary for PDI to catalyze effective protein reduction; however, placing Grx1 into the PDI multidomain structure does not enhance its already high reductase activity. Grx1 and PDI have both found mechanisms to enhance active site reactivity toward proteins, particularly in the kinetically difficult direction: Grx1 by providing a reactive glutathione mixed disulfide to supplement its oxidase activity and PDI by utilizing its multidomain structure to supplement its reductase activity. Members of the thioredoxin superfamily of redox proteins transfer reducing and oxidizing equivalents between proteins through thiol-disulfide exchange. With overall structural similarity, the classical thioredoxin and glutaredoxin members of the family all have an active site with the sequence CXXC (1Martin J.L. Structure. 1995; 3: 245-250Abstract Full Text Full Text PDF PubMed Scopus (694) Google Scholar). The two active site cysteines are able to cycle between dithiol and disulfide redox states. Despite the similarities in the active sites and a general mechanism involving thiol-disulfide exchange, thioredoxin family proteins serve a diverse set of biological functions. The family patriarch is thioredoxin, the principal function of which is to provide reducing equivalents for deoxyribonucleotide synthesis by reducing the active site disulfide of ribonucleotide reductase (2Holmgren A. Annu. Rev. Biochem. 1985; 54: 237-271Crossref PubMed Google Scholar). Glutaredoxins (Grx), 1The abbreviations used are: Grx, glutaredoxin; Grx1, glutaredoxin 1 from E. coli; PDI, protein-disulfide isomerase; SG-RNase, RNase with its cysteines in mixed disulfides with glutathione. which have a GSH binding site (3Bushweller J.H. Billeter M. Holmgren A. Wuthrich K. J. Mol. Biol. 1994; 235: 1585-1597Crossref PubMed Scopus (120) Google Scholar), serve as reductants of protein-SG mixed disulfides and also provide reducing equivalents to ribonucleotide reductase apart from their many other functions (4Holmgren A. J. Biol. Chem. 1989; 264: 13963-13966Abstract Full Text PDF PubMed Google Scholar). Protein-disulfide isomerase (PDI) and its prokaryotic counterpart, DsbA, function primarily to introduce disulfides into protein substrates during oxidative protein folding (5Bardwell J.C. McGovern K. Beckwith J. Cell. 1991; 67: 581-589Abstract Full Text PDF PubMed Scopus (830) Google Scholar, 6Solovyov A. Xiao R. Gilbert H.F. J. Biol. Chem. 2004; 279: 34095-34100Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). In thioredoxin and PDI, the more N-terminal cysteine, CXXC, of the active site is exposed to solution (7Kemmink J. Darby N.J. Dijkstra K. Nilges M. Creighton T.E. Biochemistry. 1996; 35: 7684-7691Crossref PubMed Scopus (184) Google Scholar) and acts as a nucleophile to attack substrate disulfides (8Walker K.W. Lyles M.M. Gilbert H.F. Biochemistry. 1996; 35: 1972-1980Crossref PubMed Scopus (142) Google Scholar). The second active cysteine, CXXC, is buried and constrained to react with only the N-terminal active site cysteine (7Kemmink J. Darby N.J. Dijkstra K. Nilges M. Creighton T.E. Biochemistry. 1996; 35: 7684-7691Crossref PubMed Scopus (184) Google Scholar). The reduction potentials of the active site disulfides of the thioredoxin family span an extremely large range, depending, in part, on the identity of the two XX residues and the protein context in which the CXXC sequence occurs. The reduction potential of the best oxidant, DsbA (–124 mV) differs from the reduction potential of the worst oxidant, thioredoxin (–270 mV) by more than 146 mV, corresponding to a ratio of nearly 105 in thermodynamic stability during the disulfide/dithiol equilibrium (9Aslund F. Berndt K.D. Holmgren A. J. Biol. Chem. 1997; 272: 30780-30786Abstract Full Text Full Text PDF PubMed Scopus (330) Google Scholar). Within the family, there is a strong correlation between the identity of the two intervening residues and the redox potentials (10Huber-Wunderlich M. Glockshuber R. Fold Des. 1998; 3: 161-171Abstract Full Text Full Text PDF PubMed Scopus (176) Google Scholar, 11Krause G. Lundstrom J. Barea J.L. Pueyo de la Cuesta C. Holmgren A. J. Biol. Chem. 1991; 266: 9494-9500Abstract Full Text PDF PubMed Google Scholar, 12Chivers P.T. Laboissiere M.C. Raines R.T. EMBO J. 1996; 15: 2659-2667Crossref PubMed Scopus (161) Google Scholar). Both Raines and co-workers (12Chivers P.T. Laboissiere M.C. Raines R.T. EMBO J. 1996; 15: 2659-2667Crossref PubMed Scopus (161) Google Scholar) and Creighton and co-workers (13Kortemme T. Darby N.J. Creighton T.E. Biochemistry. 1996; 35: 14503-14511Crossref PubMed Scopus (111) Google Scholar) have argued that much of the difference in reduction potential can be attributed to factors that stabilize the thiolate anion of the reduced active site. The active site cysteines have been suggested to form a thiol-thiolate hydrogen bond network, which stabilizes the more N-terminal active site thiolate in the reduced form of the proteins (14Nordstrand K. F. Holmgren A. G. Berndt K.D. PubMed Scopus Google Scholar, Holmgren A. Biochemistry. 1995; PubMed Scopus Google Scholar, A. G. Structure. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). In the of the thioredoxin family proteins their thermodynamic In its reduced thioredoxin and glutaredoxin are reductants of both protein and disulfides (2Holmgren A. Annu. Rev. Biochem. 1985; 54: 237-271Crossref PubMed Google Scholar, A. J. Biol. Chem. 1989; 264: 13963-13966Abstract Full Text PDF PubMed Google Scholar). the reductants PDI and DsbA, are also reductants the of substrate disulfide formation the active DsbA and protein-disulfide isomerase are the best A between and reactivity is not the overall mechanism of thiol-disulfide exchange, the fundamental is the all the family thiol/disulfide exchange, the is J. Chem. Scopus Google Scholar), that factors the of the thiolate disulfide also be in the for thiol-disulfide exchange. also as are to the also have been in the and M. G. Holmgren A. Mol. PubMed Scopus Google Scholar, M. C. J. M. G. J. M. Holmgren A. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, J. G. J. E. Mol. Biol. Cell. PubMed Scopus Google Scholar). Glutaredoxins catalyze the reduction of protein-SG mixed their in the reducing of the (4Holmgren A. J. Biol. Chem. 1989; 264: 13963-13966Abstract Full Text PDF PubMed Google Scholar). PDI and DsbA are in the more oxidizing of the and in with their function as Escherichia Grx1 is a PDI is a multidomain N.J. E. R. J. Mol. Biol. 1998; PubMed Scopus Google Scholar). PDI is of structural all of which a thioredoxin The two catalytic domains of PDI and are by two intervening structural domains and The multidomain structure is in the of disulfide during protein is not for PDI oxidase activity. PDI catalyzes disulfide in its principal function in is to serve as an oxidase for disulfide formation in the A. Xiao R. Gilbert H.F. J. Biol. Chem. 2004; 279: 34095-34100Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). In of the of and in the thioredoxin family, by the that Grx1 is a much better oxidase for protein than PDI the that PDI is a 600-fold better oxidant, (9Aslund F. Berndt K.D. Holmgren A. J. Biol. Chem. 1997; 272: 30780-30786Abstract Full Text Full Text PDF PubMed Scopus (330) Google Scholar). the of the of glutaredoxin as a protein oxidase and reductase, have the of disulfide formation and reduction by Grx1 and PDI. also a protein with the Grx1 structure in of one of the active site domains in PDI. of the more active site cysteines of Grx1 and PDI that Grx1, not PDI, can a active site cysteine to catalyze protein oxidation through the formation of a mixed As an Grx1 oxidizing equivalents to its substrates through a reactive mixed disulfide with glutathione. As protein-disulfide both Grx1 and PDI two active site cysteines. PDI, its multidomain structure is to catalyze protein disulfide whereas domains are not to catalyze substrate It is suggested that both Grx1 and PDI have mechanisms to enhance of that be difficult because of the thermodynamic stability of the active site and ribonuclease A and glutathione reductase from The peptide substrate used in from It the sequence with the and and in solution the of to and Biochem. 1989; PubMed Scopus Google Scholar). and with a active site cysteine (8Walker K.W. Lyles M.M. Gilbert H.F. Biochemistry. 1996; 35: 1972-1980Crossref PubMed Scopus (142) Google Scholar). of E. Grx1 to the from to The of Xiao R. A. Gilbert H.F. Holmgren A. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar) to the Grx1-PDI The an N-terminal Grx1 with the domains of PDI a The cysteines in the catalytic of PDI both to that the only catalytic from the Grx1 PDI, Grx1, and their proteins from with an N-terminal R. A. Gilbert H.F. Holmgren A. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). a as R. A. Gilbert H.F. Holmgren A. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar) and than by of RNase reduced RNase by reduction of the protein in as M. J. F. G. Biochemistry. 1997; PubMed Scopus Google Scholar). The mixed disulfide of RNase and glutathione as in J. A. F. Holmgren A. PubMed Scopus Google Scholar, RNase A as in PubMed Scopus Google exchange between Grx1, and PDI its domains in 1 and GSH with a peptide of and an of 1 of and thiol-disulfide exchange by the of to a of and reduced peptide in the and on a of in in a of the of the to to the and by their The initial velocity of the peptide oxidation from the initial of formation of the toward the of RNase activity by the of to the of RNase activity as by Lyles and Gilbert M.M. Gilbert H.F. Biochemistry. 1991; PubMed Scopus Google Scholar). A glutathione redox buffer used to the redox The of reduced RNase of the of active RNase show a during which the reduced RNase is to a of nearly disulfides M. J. F. G. Biochemistry. 1997; PubMed Scopus Google Scholar, M.M. Gilbert H.F. Biochemistry. 1991; PubMed Scopus Google Scholar). The by the in RNase activity the to active RNase The initial velocity of dithiol oxidation from the by that for the oxidation of the disulfides of is a of the initial the is to the of the oxidase (8Walker K.W. Lyles M.M. Gilbert H.F. Biochemistry. 1996; 35: 1972-1980Crossref PubMed Scopus (142) Google Scholar), and the can be The in the of as Grx1 that not isomerase a of PDI an with only a cysteine active site a of 1 In the velocity that in the of of RNase and reduction of by the formation of to the oxidation of glutathione 1 1 glutathione reductase and The by PDI for the reduction that in the of PDI The reduction of scrambled RNase in the of GSH in and 1 glutathione The oxidation of in the of Grx1 PDI, PDI The activity the of for of reduction of Grx1 by GSH by the formation of to the oxidation of with glutathione The by the in of a of The by of Grx1 with a solution of GSH and glutathione reductase all in a GSH the for the of by to a by a in with to a oxidation of only of GSH the with the in the of the for reduced by and glutathione reductase the used to Grx1 The for the of times the for Grx1 The for Grx1 reduction as a function of GSH by to as N.J. Creighton T.E. Biochemistry. 1995; PubMed Scopus Google Scholar, H.F. Biochemistry. 1989; PubMed Scopus Google Scholar). Catalysis of substrates are with both and the reactivity of the active sites on a and to protein substrates are substrates for PDI Grx1, the oxidation of a peptide are in and the for a more of are in With peptide PDI and Grx1 are both PDI requires both active site cysteines to catalyze peptide oxidation, with a dithiol mechanism of disulfide formation as with protein The individual catalytic domains of PDI are also effective with the The other domains of PDI and the potential substrate provide are not to peptide oxidase activity. Grx1 is a superior oxidase on active site with PDI. A in on the peptide oxidase that Grx1 catalyzes the peptide oxidation through a mechanism that requires only the cysteine 1 and of peptide oxidation by Grx1, PDI, and with a cysteine in the active oxidase in a Catalysis of the of the oxidative of reduced the initial of disulfide formation RNase G. M. Mol. Biol. Google Scholar). there is a the of the is with the formation of RNase disulfides that be and to and can be used as an of the of protein oxidation (8Walker K.W. Lyles M.M. Gilbert H.F. Biochemistry. 1996; 35: 1972-1980Crossref PubMed Scopus (142) Google Scholar). As can be in the of the of oxidative PDI, the with the as as the of disulfides to RNase A of Grx1, in to the PDI that is the the to the isomerase with the by and Holmgren J. Holmgren A. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar). PDI is a 600-fold better than Grx1 (9Aslund F. Berndt K.D. Holmgren A. J. Biol. Chem. 1997; 272: 30780-30786Abstract Full Text Full Text PDF PubMed Scopus (330) Google Scholar), the active site of Grx1 is a 30-fold better oxidase than PDI in a redox buffer of 1 GSH and of the oxidation of reduced RNase by Grx1, PDI, and with a cysteine in the active oxidase PDI from from from K.W. Lyles M.M. Gilbert H.F. Biochemistry. 1996; 35: 1972-1980Crossref PubMed Scopus (142) Google in a the oxidation of protein PDI catalyzes transfer of its active site disulfide into the both PDI active site cysteines are for effective of protein PDI with only the cysteine are not active and not the during the oxidative folding of reduced RNase of the active site of Grx1 to the cysteine on the of Grx1 to the in RNase The that Grx1 and oxidase that Grx1 can catalyze protein oxidation with only one of its active site cysteines and that the transfer of oxidizing equivalents from the redox buffer to the substrate through a mixed In with protein between the and the substrate protein to their as an oxidase. the catalytic domains of PDI by as effective an as the N.J. Creighton T.E. Biochemistry. 1995; PubMed Scopus Google Scholar). Grx1 a protein in protein oxidation more the N-terminal catalytic of PDI by Grx1 to a Grx1-PDI The two cysteines of the catalytic of PDI also to that the activity of Grx1 in the context of the of the PDI be As with PDI, the of the protein domains of PDI on the of Grx1 to function as an be that the of the catalytic site in the protein Grx1 to of a site from the domains of PDI, the of Grx1 and PDI to serve as on the high reactivity of an active site disulfide to introduce disulfides into protein In the of Grx1, active site disulfide is a disulfide with whereas for PDI, an disulfide between the two active site cysteines. Catalysis of Grx1 is a superior oxidase with PDI, also in the of the two proteins to catalyze In with its reduction Grx1 is a more effective reductase than PDI on a in the reduction of the protein disulfides in scrambled Grx1 requires both active site cysteines to be an effective of the more cysteine in the Grx1 active site the reductase activity by that PDI, both active site cysteines a to protein disulfide reduction J.H. F. Wuthrich K. Holmgren A. Biochemistry. PubMed Scopus Google Scholar) have found that Grx1 reduction of ribonucleotide reductase requires both active site with that of protein disulfide reduction by Grx1, in needs both active site of the reduction of scrambled RNase by PDI, Grx1, and of PDI and Grx1 with cysteines the active PDI in a that the active site that Grx1 is a 600-fold better potential more than PDI, the difference in reductase is surprisingly Darby that the individual domains of PDI by are of PDI N.J. E. R. J. Mol. Biol. 1998; PubMed Scopus Google Scholar). the multidomain structure of PDI the reductase activity the catalytic domains are in the of the that are from its substrate to of and Grx1 is a better than PDI, that Grx1 is a better than the catalytic domains of PDI, in with the 600-fold mV) difference in reduction The of a multidomain structure with a Grx1 in a chimera in which the catalytic by Grx1 The activity of the protein than Grx1 in to Grx1 to the domains of PDI does not enhance the reductase activity of The Grx1-PDI chimera is an effective of protein oxidation that the PDI of the does not the reactivity of the Grx1 active site as an oxidase. Grx1, a superior to from that be from the domains of PDI, whereas the enhance the reductase activity of the PDI. J. A. F. Holmgren A. PubMed Scopus Google Scholar), found that the Grx1, are good for protein-glutathione mixed Grx1 and PDI for their to catalyze reduction of the mixed disulfides of RNase and glutathione Grx1 is more active substrate than a catalytic of PDI the second active site cysteine to the reductase activity of Grx1 by that there be of a reduction mechanism for protein-SG mixed disulfides that a the reduction The PDI catalytic on the other its to protein-SG mixed disulfides the more cysteine is to that both active site cysteines are for for protein-SG mixed the of Grx1 for glutathione J.H. F. Wuthrich K. Holmgren A. Biochemistry. PubMed Scopus Google Scholar) an mechanism of protein reduction that only one active site of the reduction of mixed disulfides by PDI, Grx1, and with cysteines the active the activity to the in the of Grx1 the activity to the in the of Grx1 the activity to the in the of Grx1 the activity to the in the of Grx1 PDI. in a of the Grx1 of oxidation, the active site of Grx1 is reduced by the protein substrate and through oxidation by the reactivity of the Grx1 disulfide and the mixed disulfide, rapid The oxidation of glutathione to by the active site of Grx1 requires two the formation of a mixed disulfide and the of the mixed disulfide with GSH during the of oxidation and reduction by Grx1 PDI in to the for of reduction the fundamental for the of Grx1 with Grx1 mixed with of GSH in a The as a of the by the glutathione oxidation of to H.F. Biochemistry. 1989; PubMed Scopus Google Scholar). to that there be to all of the and and that the of faster than the of by Grx1 reduced by GSH and glutathione reductase to the for the in with in a for Grx1 reduction as a function of the GSH The are the and of The is to a to a The for Grx1 reduction GSH of the reduction of Grx1 is in GSH requires that the of GSH with the active site disulfide is equilibrium that the site disulfide is not to the mixed disulfide, and that the is the of the mixed disulfide with GSH PDI, GSH of there is a in the to a that is in as the of the mixed disulfide of PDI with GSH H.F. Biochemistry. 1989; PubMed Scopus Google Scholar). In the from to in GSH Grx1, a of the to the the potential for a in is not better than a the GSH that can be used the of Grx1 reduction faster than the there is in for Grx1 a on the GSH in With a on GSH the for GSH reduction of Grx1 is by the for reduction of Grx1, is to the overall equilibrium for the is the of and can be The that there is for a in the of GSH a on and on and The for the GSH reduction of the N-terminal thioredoxin of PDI are also N.J. Creighton T.E. Biochemistry. 1995; PubMed Scopus Google Scholar, H.F. Biochemistry. 1989; PubMed Scopus Google Scholar). show that the of the mixed disulfide with GSH is faster than the of mixed disulfide with The mixed disulfide is also much more reactive with GSH than the active site disulfide of PDI by a of It is that the mixed disulfide is much more reactive with GSH than the active site disulfide of Grx1 however, is by to set only a on In Grx1 and PDI reduction and oxidation by GSH and are that the during protein and peptide oxidation and reduction be the of Grx1 and PDI with the peptide protein substrates than the of the active site redox through with GSH the of peptide and protein oxidation and reduction by Grx1 and PDI of the with the peptide protein and equilibrium for the of Grx1 and PDI active sites with from second for reduction of Grx1 GSH as in in in for the formation of the Grx1 active site disulfide and GSH from the mixed are from in in in for the formation of the mixed disulfide from reduced Grx1 and are from for the overall oxidation of Grx1 by glutathione are from redox potential for the reduction of the active site disulfide an for the GSH redox of are from from N.J. Creighton T.E. Biochemistry. 1995; PubMed Scopus Google second for reduction of Grx1 GSH as in As in for the formation of the Grx1 active site disulfide and GSH from the mixed are from F. and for the formation of the mixed disulfide from reduced Grx1 and are from F. and for the overall oxidation of Grx1 by glutathione are from F. and redox potential for the reduction of the active site disulfide an for the GSH redox of are from F. and in a members of the thioredoxin family, the of the dithiol active sites the of disulfide formation the active site. The disulfides are the best whereas the are the best Grx1 is an As from its redox Grx1 is a better reductase than an individual catalytic of PDI and Grx1 is also a better oxidase than PDI and potential for the of Grx1 as an oxidase in the of the active site that is in the the its oxidase activity is In the redox used to the oxidase activity and only a of the PDI is to be in the whereas Grx1 is to be In the of the Grx1 active site that are as the disulfide and the mixed disulfide can be from the equilibrium and equilibrium with the redox buffer of the Grx1, whereas the disulfide a the high reactivity of the mixed disulfide that can be a kinetically with GSH faster than the active site disulfide of PDI. The mixed disulfide is the reactive glutathione mixed disulfide to and its thermodynamic stability to to a in a redox In to Grx1, PDI requires both active site cysteines to effective disulfide The that the velocity of substrate oxidation is than of a active site on the the PDI active site only the cysteine that disulfide formation by PDI is by a transfer of a disulfide to the substrate and that glutathione is not is by in from a in that a for protein disulfide bond formation in the PDI oxidizing equivalents from to proteins by thiol-disulfide exchange In of glutathione from the on disulfide bond formation in proteins Mol. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar, Mol. Cell. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar) the second active site cysteine in Grx1 to Grx1 and the have activity in the of RNase folding and a disulfide bond into a peptide and It been that the of Grx1 with PDI in of RNase in the of glutathione J. Holmgren A. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar). oxidation by mechanisms glutathione is necessary and only one active site cysteine is The high oxidase activity of that in to its as a reducing Grx1 also in protein oxidation during oxidative The high reactivity of the disulfide of be to the of the N-terminal in Grx1 and the to serve as a good by a protein J. K. Berndt K.D. J. Mol. Biol. PubMed Scopus Google Scholar, Biochemistry. 1991; PubMed Scopus Google Scholar, J. Biol. Chem. 1991; 266: Full Text PDF PubMed Google Scholar, 2004; Full Text PDF PubMed Google Scholar). The oxidase activity of PDI does not that be from the multidomain structure of PDI. A catalytic is as good an oxidase as PDI. reduction does the multidomain the individual catalytic domains are reduction is by between the substrate and the other domains of PDI, the of PDI to serve as a protein The function of the substrate be to of the the binding the of the reduction more mechanism in the reduction of protein substrates by PDI is not the domains of PDI to Grx1 does not enhance its oxidase reductase that the reactivity of the Grx1 active site and the of providing a mixed disulfide is for its oxidase activity. In Grx1 and PDI are members of the thioredoxin Grx1 mechanisms for protein oxidation and reduction that are not to the other family mechanisms that a mechanism and the high reactivity of a mixed mechanism for the reduction potential of the Grx1 active site and that Grx1, its reducing also catalyze protein The formation of protein-SG mixed disulfides by Grx1 through a mechanism an in more of protein particularly the redox of the more as of oxidative have been in and J. E. A. E. Mol. Cell. Biol. PubMed Scopus Google Scholar). The of that are for their and their functions be by their dithiol The biological of and are not is in the of proteins in the J. G. J. E. Mol. Biol. Cell. PubMed Scopus Google Scholar) and a in protein oxidative J. E. A. E. Mol. Cell. Biol. PubMed Scopus Google Scholar). PDI is because is a more effective than on its redox In the of reduction is by involving in substrate an activity that is to disulfide during oxidative protein folding M. Gilbert H.F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar).