Reaction participants Show >> << Hide
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Namehelp_outline
[protein]-disulfide
Identifier
RHEA-COMP:10593
Reactive part
help_outline
- Name help_outline L-cystine residue Identifier CHEBI:50058 Charge 0 Formula C6H8N2O2S2 SMILEShelp_outline C([C@@H](N*)CSSC[C@@H](C(=O)*)N*)(=O)* 2D coordinates Mol file for the small molecule Search links Involved in 51 reaction(s) Find molecules that contain or resemble this structure Find proteins in UniProtKB for this molecule
- Name help_outline glutathione Identifier CHEBI:57925 Charge -1 Formula C10H16N3O6S InChIKeyhelp_outline RWSXRVCMGQZWBV-WDSKDSINSA-M SMILEShelp_outline [NH3+][C@@H](CCC(=O)N[C@@H](CS)C(=O)NCC(=O)[O-])C(=O)[O-] 2D coordinates Mol file for the small molecule Search links Involved in 100 reaction(s) Find molecules that contain or resemble this structure Find proteins in UniProtKB for this molecule
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Namehelp_outline
[protein]-dithiol
Identifier
RHEA-COMP:10594
Reactive part
help_outline
- Name help_outline L-cysteine residue Identifier CHEBI:29950 Charge 0 Formula C3H5NOS Positionhelp_outline C1 SMILEShelp_outline C(=O)(*)[C@@H](N*)CS 2D coordinates Mol file for the small molecule Search links Involved in 123 reaction(s) Find molecules that contain or resemble this structure Find proteins in UniProtKB for this molecule
- Name help_outline L-cysteine residue Identifier CHEBI:29950 Charge 0 Formula C3H5NOS Positionhelp_outline C2 SMILEShelp_outline C(=O)(*)[C@@H](N*)CS 2D coordinates Mol file for the small molecule Search links Involved in 123 reaction(s) Find molecules that contain or resemble this structure Find proteins in UniProtKB for this molecule
- Name help_outline glutathione disulfide Identifier CHEBI:58297 Charge -2 Formula C20H30N6O12S2 InChIKeyhelp_outline YPZRWBKMTBYPTK-BJDJZHNGSA-L SMILEShelp_outline [NH3+][C@@H](CCC(=O)N[C@@H](CSSC[C@H](NC(=O)CC[C@H]([NH3+])C([O-])=O)C(=O)NCC([O-])=O)C(=O)NCC([O-])=O)C([O-])=O 2D coordinates Mol file for the small molecule Search links Involved in 36 reaction(s) Find molecules that contain or resemble this structure Find proteins in UniProtKB for this molecule
Cross-references
RHEA:21064 | RHEA:21065 | RHEA:21066 | RHEA:21067 | |
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Reaction direction help_outline | undefined | left-to-right | right-to-left | bidirectional |
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Publications
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Kinetic analysis of the mechanism of insulin degradation by glutathione-insulin transhydrogenase (thiol: protein-disulfide oxidoreductase).
Chandler M.L., Varandani P.T.
Kinetic studies have been made with glutathione-insulin transhydrogenase, an enzyme which degrades insulin by promoting cleavage of its disulfide bonds via sulfhydryl-disulfide interchange. The degradation of 125I-labeled insulin by enzyme purified from beef pancreas was studied with various thiol ... >> More
Kinetic studies have been made with glutathione-insulin transhydrogenase, an enzyme which degrades insulin by promoting cleavage of its disulfide bonds via sulfhydryl-disulfide interchange. The degradation of 125I-labeled insulin by enzyme purified from beef pancreas was studied with various thiol-containing compounds as cosubstrates. The apparent Km for insulin was found to be a function of the type and concentration of thiol; values obtained were in the range from 1 to 40 muM. Lineweaver-Burk plots for insulin as varied substrate were linear, whereas those for the thiol substrates were nonlinears: the plots for low molecular weight monothiols (GSH and mercaptoethanol) were parabolic; those for low molecular weight dithiols (dithiothreitol, dihydrolipoic acid, and 2,3-dimercaptopropanol) were apparently linear modified by substrate inhibition; and the plots for protein polythiols (reduced insulin A and B chains and reduced ribonuclease) were parabolic with superposed substrate inhibition. The nonparallel nature of the reciprocal plots for all substrates shows that the enzyme does not follow a ping-pong mechanism. Product inhibition studies were performed with GSH as thiol substrate. Oxidized glutathione was found to be a linear competitive inhibitor vs. both GSH and insulin. The S-sulfonated derivative of insulin A chain was also linearly competitive vs. both substrates. Inhibition by S-sulfonated B chain was competitive vs. insulin; the data eliminated the possibility that this derivative was uncompetitive vs. GSH. Experiments with the cysteic acid derivatives of insulin A and B chains similarly excluded the possibility that these were uncompetitive vs. either substrate. These inhibition studies indicate that the enzyme probably follows a randdom mechanism. << Less
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Di-fluoresceinthiocarbamyl-insulin: a fluorescent substrate for the assay of protein disulfide oxidoreductase activity.
Heuck A.P., Wolosiuk R.A.
We have developed a novel method for the continuous assay of protein disulfide oxidoreductase activity using as substrate bovine pancreas insulin in which both N-terminal amino groups are chemically modified with fluorescein isothiocyanate. The reduction of intercatenary disulfide bonds of di-fluo ... >> More
We have developed a novel method for the continuous assay of protein disulfide oxidoreductase activity using as substrate bovine pancreas insulin in which both N-terminal amino groups are chemically modified with fluorescein isothiocyanate. The reduction of intercatenary disulfide bonds of di-fluoresceinthiocarbamyl-insulin with dithiothreitol initially lowers but subsequently enhances the emission intensity. In this biphasic kinetics, the rate of increase is sensitive enough for the estimation of Escherichia coli thioredoxin concentrations from 5 nM (0.06 microgram/ml) to 500 nM (6 micrograms/ml). Neither changes of pH over a range of 6.2 to 8.4 nor neutral salts (K+, Mg2+, and Ca2+) at concentrations lower than 100 mM affect this simple reaction system. Moreover, the fluorometric method is functional for measuring the reductive capacity of Brassica napus protein disulfide isomerase. Hence, a highly reproducible and accurate one-state assay for protein disulfide oxidoreductase activity not only greatly improves the sensitivity compared to the commonly used turbidimetric assay but also represents a reliable alternative to assays based on accessory enzymes or radiolabeled substrates. << Less
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Functional characterization of ERp18, a new endoplasmic reticulum-located thioredoxin superfamily member.
Alanen H.I., Williamson R.A., Howard M.J., Lappi A.-K., Jaentti H.P., Rautio S.M., Kellokumpu S., Ruddock L.W.
Native disulfide bond formation in the endoplasmic reticulum is a critical process in the maturation of many secreted and outer membrane proteins. Although a large number of proteins have been implicated in this process, it is clear that our current understanding is far from complete. Here we desc ... >> More
Native disulfide bond formation in the endoplasmic reticulum is a critical process in the maturation of many secreted and outer membrane proteins. Although a large number of proteins have been implicated in this process, it is clear that our current understanding is far from complete. Here we describe the functional characterization of a new 18-kDa protein (ERp18) related to protein-disulfide isomerase. We show that ERp18 is located in the endoplasmic reticulum and that it contains a single catalytic domain with an unusual CGAC active site motif and a probable insertion between beta3 and alpha3 of the thioredoxin fold. From circular dichroism and NMR measurements, ERp18 is well structured and undergoes only a minor conformational change upon dithioldisulfide exchange in the active site. Guanidinium chloride denaturation curves indicate that the reduced form of the protein is more stable than the oxidized form, suggesting that it is involved in disulfide bond formation. Furthermore, in vitro ERp18 possesses significant peptide thiol-disulfide oxidase activity, which is dependent on the presence of both active site cysteine residues. This activity differs from that of the human PDI family in that under standard assay conditions it is limited by substrate oxidation and not by enzyme reoxidation. A putative physiological role for Erp18 in native disulfide bond formation is discussed. << Less