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)

In the current study, the ability of four protease inhibitors to suppress radiation-induced transformation of C3H/10T1/2 cells was investigated. The inhibitors tested included: (i) aprotinin (a serine protease inhibitor), (ii) N-acetyl-L-tyrosine ethyl ester (a chymotrypsin substrate and competitive inhibitor of protein degradation), (iii) carboxypeptidase inhibitor (a metallo-exopeptidase inhibitor) and (iv) Inhibitor II (a chymotrypsin/trypsin inhibitor). While none of the inhibitors were toxic to the cells at the concentrations employed, only carboxypeptidase inhibitor and inhibitor II are internalized radiation-induced transformation in a statistically significant manner. Utilizing fluorescent labeled inhibitors, we found that carboxypeptidase inhibitors and Inhibitor II are internalized by the cells. Fluorescent-labeled inhibitor could be observed in the cells within 15 min of incubation and is present in distinct intercellular vacuoles within 1 h. These results indicate that carboxypeptidase inhibitor and Inhibitor II are internalized by C3H/10T1/2 cells and thus would be able to inhibit intracellular proteases (or other enzymes) involved in the conversion of a cell to the malignant state.
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PMID:Inhibition of radiation-induced transformation of C3H/10T1/2 cells by carboxypeptidase inhibitor 1 and inhibitor II from potatoes. 246 57

The kinetic properties dextran-chymotrypsin conjugate were studied by means of low molecular weight substrates. It was found that KM, kcat and kcat/KM of dextran chymotrypsin for the hydrolysis of benzoyl-L-tyrosine-ethyl-ester did not differ substantially from those of the free enzyme. However, the data found for kcat of dextran-chymotrypsin and free chymotrypsin assayed for the hydrolysis of three tripeptidyl-p-nitroanilide D-Arg-Val-Trp-pNA, D-Arg-Val-Tyr-pNA, Z-Phe-Pro-Phe-pNA, were definitely different. The inhibition of the modified chymotrypsin with soybean trypsin inhibitor was found to be less pronounced than that with the free enzyme. The effect of potassium and magnesium salts on the inactivation of both enzymes was also studied. The effect of dextran matrix on the catalytic properties and the conformational stability of modified chymotrypsin is discussed.
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PMID:Properties of chymotrypsin bound covalently to dextran. 247 Feb 17

Probable participation of sperm protease in the acrosome reaction was investigated using several inhibitors and substrates. Among those examined, L-1-tosylamide-2-phenylethyl chloromethyl ketone (TPCK) and chymostatin, chymotrypsin inhibitors, p-nitrophenyl-p'-guanidinobenzoate (NPGB), a serine protease inhibitor, and N-benzoyl-L-tyrosine ethyl ester (BTEE), a chymotrypsin substrate, inhibited the egg jelly-induced acrosome reaction of Strongylocentrotus intermedius. TPCK and BTEE, however, did not inhibit the reaction caused by ionophores, A23187, or nigericin. To know the mechanism of inhibition by chymotrypsin inhibitors and substrates of the egg jelly-induced acrosome reaction, intracellular Ca2+ concentration [( Ca2+]i) and pH (pHi) were measured with fura-2 and 2',7'-bis (carboxy-ethyl)carboxyfluorescein (BCECF), respectively. Egg jelly caused increase of [Ca2+]i, which was depressed by BTEE. Egg jelly also caused a transient rise of pHi, which was not depressed by BTEE. In the presence of verapamil, the acrosome reaction by egg jelly was significantly inhibited concomitant with depressed increase of [Ca2+]i. The rise of pHi was not depressed by verapamil. Thus, modes of action of BTEE and of verapamil are similar to each other. Bringing these findings together, the authors present a view that a chymotrypsin-like protease of sea urchin sperm activates verapamil-sensitive Ca2+ channels, which take part in the acrosome reaction.
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PMID:Activation of Ca2+ channels during the acrosome reaction of sea urchin sperm is inhibited by inhibitors of chymotrypsin-like proteases. 255 Mar 37

The effects of citrate ion concentration and pH on the optical spectra and fluorescence decay have been measured for several tyrosine model compounds and lima bean trypsin/chymotrypsin inhibitor, a protein containing one tyrosine at position 69 and seven disulfides but no tryptophan, in order to determine the location and environment of Tyr 69. Tyrosine in the protein is protected from citrate collisional quenching, as indicated by the dynamic quenching constant 9 to 15 times smaller than those for the model peptides. Static quenching remains, with a Stern-Volmer constant of about 1.0 M-1, somewhat smaller than those of L-tyrosine, tyrosine-glutamate, and leucine-tyrosine-leucine. The elevated pKa of Tyr 69, greater than or equal to 11.6, also indicates protein protection from solvent ions. Though Coulomb repulsion of the Glu 70/citrate pair may play a role in the shielding of Tyr 69 from citrate, our measurements indicate that steric effects of the protein structure are more important. Tyrosinate emission in the protein at neutral pH is minimal.
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PMID:Spectroscopy and fluorescence quenching of tyrosine in lima bean trypsin/chymotrypsin inhibitor and model peptides. 262 88

Initial rates of peptide-bond synthesis catalyzed by poly(ethylene glycol)-modified chymotrypsin in benzene were determined using high-performance liquid chromatography. Enzymatic synthesis of N-benzoyl-L-tyrosyl-L-phenylalanine amide from N-benzoyl-L-tyrosine ethyl ester and L-phenylalanine amide was found to obey Michaelis-Menten kinetics an to be consistent with a ping-pong mechanism modified by a hydrolytic branch. The catalytic activity of modified chymotrypsin was dependent on both water concentration and type of organic solvent, the highest synthesis rate being obtained in toluene. Since the chymotrypsin specificity in the organic phase was actually altered, the enzyme's apparent kinetic parameters were determined for different substrates and compared to those obtained with other serine proteases in benzene. Both N-benzoyl-L-tyrosine ethyl ester and N-alpha-benzoyl-L-lysine methyl ester were comparable acyl donors in benzene and the (kcat/Km)app value of modified chymotrypsin was only 10-fold smaller than that obtained with poly(ethylene glycol)-modified trypsin in the synthesis of N-alpha-benzoyl-L-lysyl-L-phenylalanine amide. The change in chymotrypsin specificity was also confirmed through the binding of trypsin inhibitors in benzene. The overall results suggest that hydrophobic bonding between the enzyme and its substrate should not be taken into account during catalysis in the organic phase. In general, if hydrophobic interactions are involved in the binding of substrates to the active site in aqueous media, the replacement of water by hydrophobic solvents will induce some change in enzyme specificity. Moreover, secondary residues of enzyme-binding sites may also exert a significant influence on specificity since, as observed in this study, chymotrypsin exhibited high affinity for cationic substrates and cationic inhibitors as well in apolar solvents.
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PMID:Kinetics and specificity of serine proteases in peptide synthesis catalyzed in organic solvents. 265 20

The abilities of several nucleotides to protect tyrosine aminotransferase (L-tyrosine: 2-oxoglutarate aminotransferase, EC 2.6.1.5) against proteolytic inactivation in vitro have been examined as part of an ongoing investigation of the role of cyclic GMP in the intracellular degradation of the hepatic enzyme. Although neither cyclic GMP nor cyclic AMP was found to exert such a protective effect, certain nucleotide analogs were observed to inhibit the inactivation of tyrosine aminotransferase by trypsin and chymotrypsin. The nucleotides which conferred the strongest protection were the dibutyryl derivatives of cyclic GMP and cyclic AMP. This phenomenon appears to require a purine nucleotide with hydrophobic substituent(s), while the cyclic phosphate is not essential. The nucleotides probably act by direct interaction with tyrosine aminotransferase as indicated by changes in kinetic properties and heat stability of the enzyme and by their failure to inhibit trypsin when other protein substrates, including another aminotransferase, were used. Dibutyryl cyclic AMP was shown to block the appearance of a characteristic 43 kDa tryptic cleavage product of tyrosine aminotransferase but not the conversion of the native 54 kDa form to a size of 50 kDa. Arguments are presented against the involvement of the protective effect in the actions of dibutyryl cyclic nucleotides on tyrosine aminotransferase in cells.
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PMID:Protection of tyrosine aminotransferase against proteolytic digestion by nucleotide derivatives. 290 Jun 54

The sea urchin blastula secretes a hatching enzyme (HE) that dissolves the fertilization envelope. HE was collected from the supernatant seawater of cultures of hatched Strongylocentrotus purpuratus blastulae, and concentrated 20 times by ultrafiltration. The proteolytic activity of HE using casein as substrate was inhibited by the chymotrypsin inhibitors, chymostatin and N-tosyl-L-phenylalanine chloromethyl ketone. The activity was not inhibited by inhibitors (antipain, elastatinal, pepstatin, phosphoramidon, soybean trypsin inhibitor, and N alpha-p-tosyl-L-lysine chloromethyl ketone) of other types of proteases. HE did not hydrolyze the synthetic trypsin substrate, alpha-N-benzoyl-L-arginine ethyl ester, but did hydrolyze the synthetic substrate of chymotrypsin, N-benzoyl-L-tyrosine ethyl ester (BTEE). The BTEEase activity of HE was completely inhibited by the chymotrypsin inhibitors chymostatin and 2-nitro-4-carboxyphenyl N,N-diphenylcarbamate (NCDC). Chymostatin inhibited the natural hatching of sea urchin blastulae. Application of HE to freshly fertilized sea urchin eggs, 2 h after insemination, caused premature dispersal of the hardened fertilization envelope. Chymostatin and NCDC inhibited HE-induced lysis of the fertilization envelope, while inhibitors of other types of proteases were ineffective. These data suggest that sea urchin HE is a chymotrypsin-like protease we call "chymotrypsin."
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PMID:Evidence that hatching enzyme of the sea urchin Strongylocentrotus purpuratus is a chymotrypsin-like protease. 307 14

Chymotrypsin modified with polyethylene glycol was successfully used for peptide synthesis in organic solvents. The benzene-soluble modified enzyme readily catalyzed both aminolysis of N-benzoyl-L-tyrosine p-nitroanilide and synthesis of N-benzoyl-L-tyrosine butylamide in the presence of trace amounts of water. A quantitative reaction was obtained when either hydrophobic or bulky amides of L- as well as D-amino acids were used as acceptor nucleophiles, while almost no reaction occurred with free amino acids or ester derivatives. The acceptor nucleophile specificity of modified chymotrypsin as a catalyst in the formation of both amide and peptide bonds in organic solvents was quite comparable to that in aqueous solution as well as to that of the leaving group in hydrolysis reactions. By contrast, the substrate specificity of modified chymotrypsin in organic solvents was different from that in water since arginine and lysine esters were found to be as effective as aromatic amino acids to form the acyl-enzyme with subsequent synthesis of a peptide bond.
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PMID:Peptide synthesis catalyzed by polyethylene glycol-modified chymotrypsin in organic solvents. 339 94

A method--enzymoblotting--was developed for localizing various enzymes after electrophoretic separation, transfer to nitrocellulose, and incubation with specific substrates. As an application, the proteinases porcine trypsin (EC 3.4.21.4), bovine chymotrypsin (EC 3.4.21.1), porcine elastase (EC 3.4.22.11), and their zymogen forms from porcine pancreas homogenate were analyzed utilizing specific p-nitroanilide substrates. After agarose gel electrophoresis, transfer of the separated proteinases to a nitrocellulose membrane was performed by capillary diffusion for 30 min. After air-drying of the nitrocellulose membrane, it was incubated in the appropriate substrate solution for 60 min. N-alpha-Benzoyl-DL-arginine-para-nitroanilide HCl was used as a substrate for trypsin, N-benzoyl-L-tyrosine-para-nitroanilide and succinyl-L-phenylalanine-para-nitroanilide for chymotrypsin, and N-succinyl-L-alanyl-L-alanyl-L-alanine-para-nitroanilide for elastase. p-Nitroaniline, the product thus obtained, was diazotized with N-(1-naphthyl)ethylenediamine to a red azo dye, visible at the site of the proteinases on the nitrocellulose membrane. The results could be preserved at -18 degrees C. Zymogen forms of the pancreas proteinases were detected in a similar manner. They were converted to active proteinases in situ on the nitrocellulose membrane after preincubating the nitrocellulose membrane in the activation enzymes enteropeptidase or trypsin.
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PMID:Enzymoblotting: a method for localizing proteinases and their zymogens using para-nitroanilide substrates after agarose gel electrophoresis and transfer to nitrocellulose. 351 6

Induction of a microsomal Ca2+-dependent serine protease by hepatic tumor promoters was studied. Male F344 rats were fed a diet containing one of the following promoting agents: phenobarbital (CAS: 50-06-6), dichlorophenyltrichloroethane (CAS: 50-29-3), butylated hydroxytoluene, ethyl-alpha-chlorophenoxyisobutyrate (CAS: 128-95-0), or 17-alpha-ethynylestradiol (CAS: 57-63-6) or a nonpromoting agent, diphenylhydantoin (CAS: 57-41-0), for 1 week. By treatment with promoters, the protease activity in the microsomal fraction was increased to threefold to fivefold that of control, whereas only a slight increase of activity was found after diphenylhydantoin treatment. The Ca2+-dependent protease activity was determined with the use of N-benzoyl-L-tyrosine ethyl ester as the substrate in a medium containing 50 mM CaCl2 for its maximal activity. This protease was preferentially localized in the smooth microsomal membrane and strongly inhibited by diisopropyl phosphorofluoridate (CAS: 55-91-4), and the optimum pH of the activity was 7.8. It appears that the Ca2+-dependent serine protease measured by using a chymotrypsin substrate is a novel protease, and induction of its activity by hepatic tumor-promoting agents is a common and specific phenomenon.
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PMID:Induction of a novel Ca2+-dependent chymotrypsin-like serine protease by tumor promoters in rat livers. 352 96


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