Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.1 (chymotrypsin)
10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The proteolytic digestion of GPIIIa on intact platelets by chymotrypsin, thrombin, plasmin, trypsin, and staphylococcal V8 protease was monitored in immunoblot studies employing three different antibodies to GPIIIa, one of which was made against a 13-residue synthetic peptide containing the amino terminus of GPIIIa. Chymotrypsin, plasmin, and trypsin gave similar patterns, from which it could be inferred that the major products after extensive digestion were two-chain molecules composed of amino-terminal fragments of Mr approximately 17,000-18,000 disulfide bonded to carboxyl-terminal remnants of Mr approximately 58,000-70,000. These patterns suggest that GPIIIa contains a large disulfide-bonded loop of at least 325 amino acids that is susceptible to proteolytic cleavage, and that the 4 cysteine residues between residues 177 and 273 bond with each other. Such a structure can also account for the presence of the PIA1 epitope, which has recently been localized to a polymorphism at position 33 on these late digestion products. Thrombin did not proteolyze GPIIIa even at 2.5 units/ml. Still to be resolved is whether the minor immunoreactive GPIIIa bands found on normal platelets are related to in vivo or in vitro proteolysis and whether GPIIIa proteolysis plays a role in chymotrypsin-induced exposure of the GPIIb/IIIa receptor.
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PMID:Evidence that platelet glycoprotein IIIa has a large disulfide-bonded loop that is susceptible to proteolytic cleavage. 252 61

The functional domains of the in situ red cell membrane calcium pump were mapped by a double labeling technique. In inside-out vesicles (IOVs) the calcium pump was phosphorylated by [gamma-32P]ATP, the proteins blotted onto nitrocellulose and tagged by monoclonal antibodies raised against the purified pump protein. After proteolytic treatment of the IOVs by trypsin, chymotrypsin, or calpain-I, the fragmentation pattern of the enzyme was followed on the double-labeled immunoblots. The changes in the kinetics of the pump were examined by parallel measurements of the active calcium uptake in IOVs. By analysis of the results of tryptic digestion, it was possible to show that the antibodies recognized three different domains of the pump: 1) a Mr = 10,000-15,000 fragment (not seen directly) which includes the calmodulin-binding domain, 2) a nonphosphorylated Mr = 35,000 tryptic fragment, and 3) a phosphorylated fragment of Mr = 76,000-81,000. Chymotrypsin or calpain-I digestion of the membranes produced one major, Mr = 125,000 fragment, which had lost antibody-binding region 1. Production of this fragment coincided with the loss of calmodulin dependence and with a calmodulin-like activation of IOV calcium uptake (high Vmax, cooperativity in calcium activation). The Mr = 125,000 fragment was further activated by acidic lipids producing high Vmax and low K 1/2 (Ca2+) with no cooperativity. Based on these data a kinetic model and a functional map of the plasma membrane calcium pump is suggested.
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PMID:Functional domains of the in situ red cell membrane calcium pump revealed by proteolysis and monoclonal antibodies. Possible sites for regulation by calpain and acidic lipids. 253 49

The effect of chymotrypsin on aldosterone biosynthesis by dispersed rabbit adrenal capsular cells was examined. Bovine alpha-chymotrypsin at concentrations of 10(-7) to 10(-5) M stimulated aldosterone production, and human chymotrypsin had an even stronger stimulatory effect. Bovine trypsin had no effect on aldosterone production by adrenal cells. Chymotrypsin treatment did not change the sensitivity of the adrenal cells to ACTH or angiotensin II. These results suggest the existence of unique chymotrypsin-susceptible sites on rabbit adrenal capsular cells, the digestion of which results in stimulation of aldosterone biosynthesis.
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PMID:Proteolytic activation of aldosterone biosynthesis in rabbit adrenal capsular cells by alpha-chymotrypsin. 255 36

A fluorescent peptide substrate to explore the protease specificity for the amino acid regions C- and N-terminal to the cleavage site has been designed. Intramolecular quenching of indole fluorescence by an N-terminal dansyl group separated by six amino acid residues forms the basis of this assay. For a particular enzyme, specificity can be designed into the peptide sequence by means of the number of residues that separate the two chromophores. In the present instance, the heptapeptide Dns-Gly-Lys-Tyr-Ala-Pro-Trp-Val is used to assay angiotensin converting enzyme (ACE), Astacus protease, carboxypeptidase A, alpha-chymotrypsin, and trypsin, all of which cleave the peptide in accord with their known specificity: Trypsin and Astacus protease hydrolyze only the Lys-Tyr and Tyr-Ala bonds, respectively. alpha-Chymotrypsin primarily cleaves the Tyr-Ala bond while ACE makes three successive dipeptidyl cleavages from the C-terminus. Carboxypeptidase rapidly hydrolyzes first the Trp-Val and then the Pro-Trp bond. For all of the enzymes, catalytic activity (kcat/Km) is in the range from 10(5) to 10(6) M-1 s-1. Hydrolysis causes a fluorescence increase in the 310 to 410 nm region of 8.6- to 13.6-fold depending on the enzyme that is assayed. Assays can be designed based on the increase in tryptophan fluorescence or by individual product analyses using thin-layer or high-performance liquid chromatography. The specificity and sensitivity of such internally quenched fluorescent oligopeptides would seem to be ideal for the assay of specific endoproteases.
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PMID:A fluorescent oligopeptide energy transfer assay with broad applications for neutral proteases. 255 28

A highly branched polyethyleneimine (PEI) was used as a spacer for immobilizing alpha-chymotrypsin on the surface of Langmuir-Blodgett (LB) membranes which were deposited on the gate of an ion-sensitive field effect transistor (ISFET). alpha-Chymotrypsin could be covalently immobilized through the glutaraldehyde-activated PEI on the LB membrane-coated ISFET. The alpha-chymotrypsin-modified ISFET showed a potentiometric response to the substrate at concentrations of more than 0.1 mM. Some performance characteristics of the sensor, such as pH response, response time, and long-term stability were examined.
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PMID:Enzyme sensors based on an ion-sensitive field effect transistor coated with Langmuir-Blodgett membranes. Use of polyethyleneimine as a spacer for immobilizing alpha-chymotrypsin. 263 78

A new method for the determination of the artificial sweetener aspartame is described. alpha-Chymotrypsin is used to cleave the methyl ester group of aspartame, producing methanol hydrolytically. The methanol is detected using an electrode which is constructed by physically trapping yeast alcohol oxidase enzyme at the tip of a dissolved oxygen electrode. The decrease in oxygen concentration, which occurs as methanol is enzymatically oxidized to formaldehyde, is measured amperometrically. Aspartame levels in diet soft drinks as determined by the proposed method and by liquid chromatography are in excellent agreement. The relative standard deviation of the measurements is 0.83%. The methanol present in diet cola as a result of aspartame degradation can also be measured by using the electrode without alpha-chymotrypsin.
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PMID:Determination of aspartame in beverages using an alcohol oxidase enzyme electrode. 265 24

1. The effects of peptidase enzymes on non-adrenergic, non-cholinergic (NANC) inhibitory responses of guinea-pig trachea to electrical field stimulation (EFS), and on relaxations induced by vasoactive intestinal peptide (VIP) and peptide histidine isoleucine (PHI) have been examined. 2. alpha-Chymotrypsin reduced both the magnitude and, particularly, the duration of the inhibitory response to EFS, whereas papain reduced only the magnitude. Aprotinin, a peptidase inhibitor prevented the effects of alpha-chymotrypsin but was without effect on papain. 3. alpha-Chymotrypsin and papain both abolished relaxant responses to exogenous VIP and PHI. The action of alpha-chymotrypsin was prevented by aprotinin, whereas that of papain was not affected. 4. The peptidases were without effect on concentration-response curves to methacholine or to isoprenaline. It was also observed that, in the absence of the peptidases, aprotinin had no effect on inhibitory responses either to EFS or to exogenous VIP and PHI. 5. It is suggested that neuropeptides, possibly VIP and PHI, released during EFS of guinea-pig trachea, partly mediate NANC relaxations, and that their action may be inhibited by peptidases. However, the lack of effect of aprotinin alone, on responses to EFS, suggests that, if endogenous peptidases are important in terminating the action of neuropeptides, they are resistant to the effect of this particular peptidase inhibitor. It is further suggested that neurogenic relaxation of guinea-pig trachea is also partly mediated by a substance, possibly non-peptide, other than VIP or PHI.
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PMID:Effects of peptidases on non-adrenergic, non-cholinergic inhibitory responses of tracheal smooth muscle: a comparison with effects on VIP- and PHI-induced relaxation. 265 4

We investigated whether big endothelin (porcine 1-40) had contractile activity in isolated rat aorta or pressor activity when injected intravenously into the anesthetized rat. When isolated rat aorta was exposed to a 100 nM concentration of big endothelin, 4.8% of a maximal KCl contraction was observed, compared to 131% of KClmax when paired aortic rings were exposed to an equivalent concentration of synthetic endothelin. Likewise, big endothelin had very weak pressor activity when injected intravenously into anesthetized, ganglion-blocked rats at 10 nmol/kg. When big endothelin was incubated with chymotrypsin, native endothelin and other peptide fragments were formed. Chymotrypsin-treated big endothelin produced an endothelin-like contraction when applied to isolated rat aortic rings, and a characteristic endothelin-like effect on blood pressure in vivo. Our results indicate that the biological activity of endothelin could be effectively blocked by inhibiting the enzyme which converts big endothelin to endothelin.
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PMID:In vitro and in vivo activity of chymotrypsin-activated big endothelin (porcine 1-40). 266 Jul 86

Fecal chymotrypsin (FCT) has been measured by a new photometric method (Monotest Chymotrypsin; Boehringer, Mannheim) in 78 patients: 44 with chronic pancreatitis and 34 not affected by any pancreatic disease. The results were compared with those from other tests of pancreatic secretory (secretin-cerulein test) and digestive [serum and urinary p-aminobenzoic acid (PABA) and pancreolauryl] capacity. When FCT values were severely reduced (below 6.7 U/g), from 90 to 100% of the patients also presented abnormal pancreatic secretory and digestive capacity. On the other hand, 87% of the patients with normal FCT (above 20 U/g) presented normal secretory and digestive capacity. Patients with intermediate FCT values (between 6.7 and 20 U/g) showed normal or abnormal pancreatic secretory and digestive capacity with the same probability. Therefore, FCT, carried out as a first test, seems to identify subjects that need no further pancreatic function tests (normal and severely impaired FCT) and patients who need other more complex functional investigations (intermediate FCT values).
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PMID:The fecal chymotrypsin photometric assay in the evaluation of exocrine pancreatic capacity. Comparison with other direct and indirect pancreatic function tests. 273 75

The polypeptide composition of unfertilized, fertilized, and protease-treated zona-free mouse eggs was evaluated in this study. Zona-free eggs were radioiodinated by an Iodogen-catalyzed reaction. Light microscopic autoradiography of egg sections revealed that labeling was restricted to the cell surface. Labeled eggs were solubilized, and cell surface polypeptides were identified by one-dimensional SDS polyacrylamide gel electrophoresis and autoradiography. The unfertilized egg demonstrated 8-10 peptides that incorporated 125I, with major bands observed at approximately 145-150, 94, and 23 kilodaltons (kD). Zona-free eggs fertilized in vitro and then radiolabeled demonstrated several new bands in comparison to unfertilized eggs, with a major band appearing at approximately 36 kD. Treatment of radiolabeled unfertilized eggs with either trypsin or chymotrypsin (1 mg/ml for 5-20 min) caused enzyme-specific modifications in labeled polypeptides. Trypsin (T) treatment resulted in time-dependant modification of the three major peptides at 145-150, 94, and 23 kD. Chymotrypsin (CT) treatment, in contrast, was associated with loss or modification of the 94 kD band, with no apparent effect on either the 145-150 or 23 kD band. Taken together with previous data indicating that T or CT egg treatment interferes with sperm-egg attachment and fusion (Boldt et al.: Biol Reprod 39:19-27, 1988), these results suggest a possible role for the 94 kD protein in sperm-egg interaction.
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PMID:Characterization of cell surface polypeptides of unfertilized, fertilized, and protease-treated zona-free mouse eggs. 274 6


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