Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.27 (thermolysin)
 1,894 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A glutaredoxin was purified from rabbit bone marrow, and its amino acid sequence was determined by high performance tandem mass spectrometry. The sequences of peptides generated by digestion with trypsin alone or in combination with thermolysin were determined from their collision-induced dissociation (CID) mass spectra. Alignment of these sequences and additional sequence information were obtained from the collision-induced dissociation mass spectra of peptides obtained from digestion of the intact protein with Staphylococcus aureus V8 protease and alpha-chymotrypsin. The resulting sequence of 106 amino acids is as follows: Ac-Ala-Gln-Glu-Phe-Val-Asn-Ser-Lys-Ile-Gln-Pro-Gly-Lys-Val-Val-Val-Phe- Ile-Lys-Pro-Thr-Cys-Pro-Tyr-Cys-Arg-Lys-Thr-Gln-Glu-Ile-Leu-Ser-Glu-Leu- Pro-Phe - Lys-Gln-Gly-Leu-Leu-Glu-Phe- Val-Asp-Ile-Thr-Ala-Thr-Ser-Asp-Met-Ser-Glu-Ile- Gln-Asp-Tyr-Leu-Gln-Gln-Leu-Thr-Gly-Ala-Arg- Thr-Val-Pro-Arg-Val-Phe-Leu-Gly-Lys-Asp-Cys-Ile- Gly-Gly-Cys-Ser-Asp-Leu-Ile-Ala-Met-Gln-Glu-Lys- Gly-Glu-Leu-Leu-Ala-Arg-Leu-Lys-Glu-Met-Gly- Ala-Leu-Arg-Gln. This glutaredoxin strongly resembles the corresponding calf and pig proteins (known as glutaredoxin and thioltransferase, respectively) with respect to its primary structure and enzymatic activity as a GSH:disulfide thioltransferase, an activity also found for the glutaredoxin from Escherichia coli. However, rabbit glutaredoxin was not active as a hydrogen donor for the reduction of ribonucleotides in the presence of the ribonucleotide reductases from rabbit bone marrow, Lactobacillus leichmannii, and Corynebacterium nephridii.
 ...
PMID:Glutaredoxin from rabbit bone marrow. Purification, characterization, and amino acid sequence determined by tandem mass spectrometry. 268 77

Glutathione-insulin transhydrogenase (EC 1.8.4.2) catalyzes the inactivation of insulin through scission of the disulfide bonds to form insulin A and B chains. In the liver, the transhydrogenase occurs primarily in the microsomal fraction where most of the enzyme is present in a latent ('inactive') state. We have isolated rat hepatic microsomes with latent transhydrogenase activity being an integral part of the vesicles. We have used these vesicles to study the topological location of glutathione-insulin transhydrogenase by investigating the effects of detergents (Triton X-100 and sodium deoxycholate), phospholipase A2 and proteinases (trypsin and thermolysin) on the latent enzyme activity. Treatment of intact vesicles with variable concentrations of detergents and phospholipase A2 resulted in the unmasking of latent transhydrogenase activity. The extent of unmasking of transhydrogenase activity is dependent upon the concentration of detergent or phospholipase used and is accompanied by a parallel release of the enzyme into the soluble fraction. Activation of the transhydrogenase by phospholipase A2 is partially inhibited by bovine serum albumin and the extent of inhibition is inversely proportional to the phospholipase concentration. In intact vesicles, latent transhydrogenase activity is resistant to proteolytic inactivation by both trypsin and thermolysin, while in semipermeable and permeable vesicles these proteases inactivate 60 and 25% of the total transhydrogenase activity, respectively. Together these results indicate that in microsomes transhydrogenase is probably weakly bound to membrane phospholipid components and that most of the enzyme is present on the cisternal surface (i.e., the luminal surface of the endoplasmic reticulum) of microsomes. Each detergent and phospholipase apparently unmasks glutathione-insulin transhydrogenase activity through disruption of the phospholipid-enzyme interaction followed by translocation of the enzyme to the soluble (cytoplasmic) fraction and not through increases in substrate availability.
 ...
PMID:Topology of glutathione-insulin transhydrogenase in rat liver microsomes. 687 Nov 88

The determination of the disulfide pairings of SETI-II, a trypsin inhibitor isolated from Sechium edule, is described herein. The inhibitor contains 31 amino acid residues per mol, 6 of which are cysteine. Forty-five nmol (160 microg) of SETI-II was hydrolyzed with 20 microg thermolysin for 48 hr at 45 degrees C, and peptides were separated by reverse phase high performance liquid chromatography (RP-HPLC). The major products were identified by amino acid composition, Edman degradation, and on the basis of the sequence of the inhibitor. The disulfide bridge pairings and (yields) are: Cys1-Cys4 (79%), Cys2-Cys5 (21%) and Cys3-Cys6 (43%). When the reduced inhibitor was reoxidized with glutathione reduced form (GSH)/glutathione oxidized form (GSSG) at pH 8.5 for 3 hr, full activity was recovered. These data show that disulfide bridge pairing and oxidation can be determined at nanomole levels and that sensitive and quantitative Edman degradation can eliminate the final time- and material-consuming step of disulfide determinations by eliminating the need to purify and cleave each peptide containing a disulfide bridge.
 ...
PMID:Determination and reoxidation of the disulfide bridges of a squash-type trypsin inhibitor from Sechium edule seeds. 1532 86