Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.1 (RNase)
 16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Thioredoxin (Trx) from Escherichia coli was compared with bovine protein disulfide-isomerase (PDI) for its ability to catalyze native disulfide formation in either reduced or randomly oxidized (scrambled) ribonuclease A (RNase). On a molar basis, a 100-fold higher concentration of Trx than of PDI was required to give the same rate of native disulfide formation measured as recovery of RNase activity. A Pro-34 to His (P34H Trx) mutation in the active site of E. coli Trx (WCGPC), mimicking the two suggested active sites in PDI (WCGHC), increased the catalytic activity in disulfide formation about 10-fold. The mutant P34H Trx displayed a 35-mV higher redox potential (E'0) of the active site disulfide/dithiol relative to wild type Trx, making it more similar to the redox potential observed for PDI. This higher redox potential correlates well with the enhanced activity and suggests a role for the histidine side chain. Enzymatic isomerization of disulfides in scrambled, oxidized RNase requires the presence of a catalytic thiol such as GSH to initiate the thiol-disulfide interchange. Bovine thioredoxin reductase, together with NADPH, could replace GSH. For oxidative folding of reduced RNase in air with Trx, P34H Trx, or PDI, catalytic amounts of sodium selenite (1 microM) resulted in rapid disulfide formation and high yields of ribonuclease activity equivalent to previously known redox buffers of GSH and GSSG. These results demonstrate no obligatory role for glutathione in disulfide formation. A possible mechanism for the unknown thiol oxidative process accompanying folding and protein disulfide formation in vivo is discussed.
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PMID:A Pro to His mutation in active site of thioredoxin increases its disulfide-isomerase activity 10-fold. New refolding systems for reduced or randomly oxidized ribonuclease. 157 42

It has been proposed that dithiol-disulfide interchange and oxidation-reduction reactions may play a role in hormone-induced receptor activation. Inspection of the sequences of the gonadotropic hormones revealed a homologous tetrapeptide (Cys-Gly-Pro-Cys) between the beta subunit of lutropin (LH) and the active site of thioredoxin (TD). The beta subunit of follitropin (FSH) has a similar sequence (Cys-Gly-Lys-Cys). Thioredoxin is a ubiquitous protein serving as an electron donor for ribonucleotide reductase, but it also exhibits disulfide isomerase activity. The catalytic activity of TD was assayed by its ability to reactivate reduced and denatured ribonuclease. In this assay, the purified ovine FSH and bovine LH preparations tested were approximately 60 and approximately 300 times, respectively, as active as TD on a molar basis. This heretofore unsuspected catalytic property of FSH and LH may be important in understanding their mechanism of receptor activation and signal transduction.
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PMID:Evidence for a novel thioredoxin-like catalytic property of gonadotropic hormones. 210 78

Thioredoxin, a known catalyst for reducing protein disulfides, was shown to catalyze efficiently the refolding of pancreatic RNase either from the reduced, denatured form or from the scrambled form containing oxidized but incorrectly paired disulfides. Thioredoxin was 1000-fold more efficient on a molar basis than the model dithiol, dithiothreitol, in reactivating reduced, denatured RNase, suggesting that thioredoxin acts as an efficient catalyst for disulfide interchange. Starting with reduced, denatured RNase, enzyme activity was recovered quantitatively with a t1/2 of 30 hr with 100 microM thioredoxin compared to only a 10-20% recovery of activity in the control using air oxidation. Oxygen further stimulated the effectiveness of thioredoxin severalfold. Thioredoxin was most effective in reactivating inactive scrambled RNase, which contained mispaired disulfides, showing a t1/2 of 2 hr. Reduced thioredoxin was optimal for catalyzing disulfide interchange in scrambled RNase, whereas oxidized thioredoxin was required for reactivation of the reduced, denatured species. Optimal reactivation of scrambled RNase required a mixture of reduced and oxidized thioredoxin. Addition of reduced thioredoxin after initiating refolding of reduced denatured RNase with oxidized glutathione effected a rapid reactivation of RNase, suggesting a two-step model for protein refolding in which the monothiol catalyzes the rapid initial formation of protein disulfides and thioredoxin catalyzes the second step of disulfide interchange. Arguments are presented suggesting that thioredoxin may serve an in vivo role analogous to the protein disulfide-isomerase (EC 5.3.4.1).
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PMID:Thioredoxin-catalyzed refolding of disulfide-containing proteins. 346 91