Kinetics and equilibria of thiol/disulfide interchange reactions of selected biological thiols and related molecules with oxidized glutathione

Keire, David A.; Strauss, Erin; Guo, Wei; Noszál, Béla; Rabenstein, Dallas L.

doi:10.1021/jo00027a023

Cited by 125 publications

(112 citation statements)

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“…The standard redox potentials of cysteamine and glutathione have been reported to be nearly the same. 16 As a result, the redox potential of both types of redox buffers did not vary in the conditions used in the experiments reported here. Given that the standard redox potential of glutathione is between 0.205 V23 and 0.26 VIO, the redox potential of the buffers used in the experiments reported here was calculated to be between -145 and -200 mV.…”

Section: Methodsmentioning

confidence: 77%

Redox conditions for stimulation of in vitro folding and assembly of the glycoprotein hormone chorionic gonadotropin

Huth

Feng

Ruddon

1994

Biotech & Bioengineering

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The formation of native disulfide bonds during in vitro protein folding can be limiting in obtaining biologically active proteins. Thus, optimization of redox conditions can be critical in maximizing the yield of renatured, recombinant proteins. We have employed a folding model, that of the beta subunit of human chorionic gonadotropin (hCG- beta), to investigate in vitro oxidation conditions that facilitate the folding of this protein, and have compared the in vitro rates obtained with the rate of folding that has been observed in intact cells. Two steps in the folding pathway of hCG-beta were investigated: the rate-limiting events in the folding of this protein, and the assembly of hCG-beta with, hCG-alpha. The rates of these folding events were determined with and without protein disulfide isomerase (PDI) using two different types of redox reagents: cysteamine and its oxidized equivalent, cystamine, and reduced and oxidized glutathione. Rates of the rate-limiting folding events were twofold faster in cysteamine/cystamine redox buffers than in glutathione buffers in the absence of PDI. Optimal conditions for hCG-beta folding were attained in a 2 mM glutathione buffer, pH 7.4, that contained 1 mg/mL PDI and in 10 microM cysteamine/cystamine, pH 8.7, without PDI. Under these conditions, the half-time of the ratelimiting folding event was 16 to 20 min and approached the rate observed in intact cells (4 to 5 min). Moreover, folding of the beta subunit under these conditions yields a functional protein, based on its ability to assemble with the alpha subunit. The rates of assembly of hCG-beta with hCG-alpha in the cysteamine/cystamine or glutathione/PDI redox buffers were comparable (t(1/2/sb> = 9 to 12 min)). These studies show that rates of folding and assembly events that involve disulfide bond formation can be optimized by a simple buffer system composed of cysteamine and cystamine.

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Section: Methodsmentioning

confidence: 77%

Redox conditions for stimulation of in vitro folding and assembly of the glycoprotein hormone chorionic gonadotropin

Huth

Feng

Ruddon

1994

Biotech & Bioengineering

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“…B, GSSG as electrophile. pH-independent rate constants for alkyl thiols (blue triangles (45,61,63)) and H 2 S (black circle, this work). Slope (␤ nuc ) is 0.36.…”

Section: Figure 5 Comparison Of the Reactivities Of Hsmentioning

confidence: 85%

Reaction of Hydrogen Sulfide with Disulfide and Sulfenic Acid to Form the Strongly Nucleophilic Persulfide

Cuevasanta

Lange

Bonanata

et al. 2015

Journal of Biological Chemistry

284

344

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Background:Hydrogen sulfide (H 2 S) modulates physiological processes in mammals. Results: The reactivity of H 2 S toward disulfides (RSSR) and albumin sulfenic acid (RSOH) to form persulfides (RSSH) was assessed. Conclusion: H 2 S is less reactive than thiols. Persulfides have enhanced nucleophilicity. Significance: This kinetic study helps rationalize the contribution of the reactions with oxidized thiol derivatives to H 2 S biology.

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“…The fact that large concentrations of reducing agents are required to affect CFTR channel gating indicates that the sulfhydryls being reduced have a very high redox potential and could potentially be oxidized at physiological concentrations of reduced glutathione. In addition, the ratio of oxidized to reduced forms of biological thiols is very sensitive to intracellular redox potential, and small changes can lead to significant alterations in the ratio of oxidized to reduced forms (42).…”

Section: Discussionmentioning

confidence: 99%

Redox Reagents and Divalent Cations Alter the Kinetics of Cystic Fibrosis Transmembrane Conductance Regulator Channel Gating

Harrington¹,

Gunderson²,

Kopito³

1999

Journal of Biological Chemistry

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Gating of the cystic fibrosis Cl؊ channel requires hydrolysis of ATP by its nucleotide binding folds, but how this process controls the kinetics of channel gating is poorly understood. In the present work we show that the kinetics of channel gating and presumably the rate of ATP hydrolysis depends on the species of divalent cation present and the oxidation state of the protein.With Ca 2؉ as the dominant divalent cation instead of Mg 2؉ , the open burst duration of the channel is increased approximately 20-fold, and this change is reversible upon washout of Ca 2؉ . In contrast, "soft" divalent cations such as Cd 2؉ interact covalently with cystic fibrosis transmembrane conductance regulator (CFTR). These metals decrease both opening and closing rates of the channel, and the effects are not reversed by washout. Oxidation of CFTR channels with a variety of oxidants resulted in a similar slowing of channel gating. In contrast, reducing agents had the opposite effect, increasing both opening and closing rates of the channel. In cell-attached patches, CFTR channels exhibit both oxidized and reduced types of gating, raising the possibility that regulation of the redox state of the channel may be a physiological mode of control of CFTR channel activity.The cystic fibrosis transmembrane conductance regulator (CFTR) 1 belongs to the superfamily of ATP-binding cassette transporters, members of which are characterized structurally by two ATP-binding motifs and functionally by their ability to couple ATP hydrolysis to transport of a substrate molecule across a membrane (1, 2). Although a role for CFTR in solute transport has been proposed (3-6), no substrate has yet been identified. CFTR remains unique among ATP-binding cassette transporters in that it couples ATP hydrolysis to the opening of a Cl Ϫ channel (6, 7), reviewed in (8). The role of ATP hydrolysis in the CFTR gating process was inferred initially from studies of CFTR gating in electrophysiological experiments using poorly hydrolyzable ATP analogs and transition state phosphate analogs (9 -12). Direct measurement of ATPase activity in purified reconstituted CFTR has confirmed that it functions as an ATPase, and the measured turnover rate of the phosphorylated form (0.5-1 s Ϫ1 ) is consistent with a model in which opening and closing of the CFTR channel gate is coupled directly to ATP hydrolysis (13).The role of ATP hydrolysis by the two nucleotide-binding folds (NBF1 and NBF2) in mediating channel gating has been elucidated by studies examining CFTR channels individually mutated in each of the NBFs (14 -16). These papers presented models in which two ATP hydrolytic events, one occurring at NBF1 and one at NBF2, are involved first in opening and then closing the CFTR channel. In one model a single cycle of ATP hydrolysis occurs at NBF1 to directly open the channel (8,14,16,17). Alternatively, a second model proposes that ATP hydrolysis at NBF1 converts the channel into a gatable state from which nucleotide binding at NBF2 can then open the channel (15). In both mod...

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Kinetics and equilibria of thiol/disulfide interchange reactions of selected biological thiols and related molecules with oxidized glutathione

Cited by 125 publications

References 0 publications

Redox conditions for stimulation of in vitro folding and assembly of the glycoprotein hormone chorionic gonadotropin

Redox conditions for stimulation of in vitro folding and assembly of the glycoprotein hormone chorionic gonadotropin

Reaction of Hydrogen Sulfide with Disulfide and Sulfenic Acid to Form the Strongly Nucleophilic Persulfide

Redox Reagents and Divalent Cations Alter the Kinetics of Cystic Fibrosis Transmembrane Conductance Regulator Channel Gating

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