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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)

An absorbent for the affinity chromatography of trypsin [EC 3.4.21.4] (AP Sepharose) was prepared. The ligand was a mixture of oligopeptides (mainly di- and tripeptides) containing L-arginine as carboxyl termini, and was obtained from a tryptic digest of protamine. Trypsin was absorbed at relatively low pH (7-4), but was not absorbed at the optimum pH of catalysis (8.2). This was clearly explained on the basis of the pH dependence of the interaction of trypsin with its products. Inactivated trypsin, trypsinogen, and chymotrypsin were not absorbed. The absorption of active trypsin was interferred with by either benzamidine or urea. From these observations, it is evident that AP Sepharose is an affinity adsorbent. AP Sepharose was useful for purification of commercial bovine trypsin. A preliminary application for the purification of Streptomyces griseus trypsin was also successful.
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PMID:Affinity chromatography of trypsin and related enzymes. I. Preparation and characteristics of an affinity adsorbent containing tryptic peptides from protamine as ligands. 0 82

Chymotrypsin is specifically adsorbed at low ionic strength and alkaline pH to hydroxyalkyl methacrylate gels with N-benzyloxycarbonylglycl-D-phenylalanine or N-benzyloxycarbonylglycyl-D-leucine attached through 1,6-hexanediamine. Chymotrypsin is not adsorbed either to the unmodified gel (Spheron) or to the gel with attached, 1,6-hexanediamine (NH2-Spheron). The adsorption of chymotrypsin to Z-Gly-D-Phe-NH2-Spheron was investigated as a function of pH and ionic strength. Trypsin is not adsorbed to this gel. Chymotrypsin isolated from a crude pancreatic extract by affinity chromatography on Z-Gly-D-Phe-NH2-Spheron had the same activity as the enzyme isolated on a column of Spheron, to which the naturally-occurring trypsin inhibitor had been coupled.
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PMID:Affinity chromatography on hydroxyalkyl methacrylate gels. III. Adsorption of chymotrypsin to poly(hydroxyalkyl methacrylates) with covalently bound benzyloxycarbonyl-glycyl-D-phenylalanine and -D-leucine as function of pH and ionic strength. 0 31

Tosyl-triethylenetetramine-Sepharose (Tos-T-Sepharose) and carbenzoxytriethylenetetramine-Sepharose (Z-T-Sepharose) were found to be adsorbents utilizable in the purification of several microbial and animal proteases. The former Sepharose derivative adsorbed alpha-chymotrypsin, trypsin, subtilisin, thermolysin and neutral subtilopeptidase at neutral pH range, and acid proteases such as pepsin and Rhizopus niveus protease at pH 3.5-6.5. alpha-Chymotrypsin and trypsin were eluted with 0.1 N acetic acid and Rhizopus protease with 0.5 N acetic acid, thermolysin with 1 M guanidine-HCl or 33% ethyleneglycol, whilst pepsin was recovered by elution with 2 M guanidine-HCl at pH 3.5. The binding of neutral subtilopeptidase and subtilisin to this adsorbent was comparatively weak and both the enzymes were recovered by elution with 0.5 M NaCl at neutral pH. On the other hand, Z-T-Sepharose was found to bind tightly to these proteolytic enzymes except neutral subtilopeptidase. Trypsin and alpha-chymotrypsin were released from the adsorbent column with 1 M p-toluenesulfonate, and subtilisin with 1 M guanidine-HCl or 33% ethyleneglycol at neutral pH region. By these chromatographic procedures, the specific activities of these proteolytic enzymes increased effectively. Comparison of the binding abilities of acetyl-, benzoyl-, tosyl- and carbobenzoxy-T-Sepharoses to these enzymes suggests that hydrophobicity of tosyl and carbobenzoxy groups plays an important role in the enzyme-adsorbent interaction.
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PMID:Purification of several proteolytic enzymes by tosyl- and carbobenzoxy-triethylene-tetramine-sepharoses. 1 98

The G-200 flow-through fraction of the extract of sea urchin eggs contained a complex form of glutathione reductase (GR) [EC 1.6.4.2]. The complex was unstable and gradually dissociated with ain increase in GR activity. The activation was facilitated by high concentrations of EDTA, KCI or (NH4)2SO4. The rate of activation by salts was apparently dependent on the ionic strength. The complex form was also activated rather quickly by treatment with proteinases such as trypsin [EC 3.4.21.4], alpha-chymotrypsin [EC 3.4.21.1] or subtilisin [EC 3.4.21.14]. Trypsin caused the complex to release the free form of GR. Even after trypsin treatment, little change was observed in the dependence of the GR activity on GSSG or NADPH concentration. The GR activity of the complex form was not inhibited at all by 0.2 mM N-ethylmaleimide (NEM) in the presence of GSSG, but was reduced to 3% in the presence of NADPH. When excess NEM was sequestered with GSH, the NEM-treated complex form was strikingly activated by trypsin, while no activation was detected with the free form of enzyme pretreated with NEM. These results suggest that the active site of GR in the complex form is largely masked by a polypeptide moiety of theinhbitiory component.
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PMID:Glutathione reductase in the sea urchin egg. III. Activation of the complex form by proteinases. 1 74

[3H]Diazepam appears to bind specifically to a single, saturable, binding site located on rat brain membranes, with an affinity constant near 3 nM at pH 7.4. Specific binding constitutes more than 90% of total binding at 0 degrees and less than 10% of total binding at 37 degrees. Arrhenius plots suggest a sharp conformational change in the diazepam receptor near 18 degrees. Mitochondrial fractions from rat kidney, liver, and lung exhibit some [3H]diazepam binding that can be displaced by nonradioactive diazepam and several other benzodiazepines. However, Ro-4864, which is almost inactive in displacing [3H]diazepam from brain membranes, is extremely potent in displacing it from kidney mitochondria. Conversely, clonazepam, the most potent inhibitor of brain binding, is an extremely weak inhibitor of kidney binding. Furthermore, diazepam binding to kidney mitochondria has an affinity constantof 40 nM, about 15 times higher than that in brain. No specific diazepam binding was detected in intestine or skeletal muscle. Thus, specific [3H]diazepam binding to membranes appears to be restricted to brain, where it is unevenly distributed: the density of diazepam receptors is about five times higher in cortex (the highest density) than in pons-meddula (lowest density). Trypsin and chymotrypsin completely abolished specific [3H]diazepambinding in brain and kidney.
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PMID:Specific benzodiazepine receptors in rat brain characterized by high-affinity (3H)diazepam binding. 2 Jun 32

Some properties of protein inhibitor for trypsin (TI) from Act. janthinus 118 were studied. It was shown that TI has an antitrypsin activity within a wide pH range with a maximum at about 9,5. At 4 degrees and 20 degrees C TI is stable for 24 hours within the pH range of 6,0--11,0. At 100 degrees C TI is more stable in the slightly acid region of pH than at neutral or alkaline conditions. Trypsin and chymotrypsin inactivate the inhibitor for 8 hours. TI inhibits trypsin, fibrinolysin, subtilisin, pronase and terrilytin, but have no effect on chymotrypsin, thrombin, papain and pepsin. The dissociation constants for the trypsin-inhibitor complex were found to be 1,7.10-8 M, 4,1.10-9 M and 2,4.10-10 M, with casein, p-nitroanilide benzoylarginine and tosylarginine methyl ester used as substrates, respectively. The corresponding dissociation rate constants for the subtilisin-inhibitor complex were equal to 1.10-9 M and 4.10-10 M with casein and carbobenzoxy-L-alanyl-L-alanyl-L-leucin p-nitroanilide used as substrates, respectively.
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PMID:[Stability and specificity of extracellular protein inhibitor for trypsin from Actinomyces janthinus 118]. 3 28

Highly sensitive gelatin substrate films prepared according to a recent variant of the procedure are studied for their susceptibility to the action of various endopeptidases and exopeptidases. Trypsin, papain, elastase, and chymotrypsin are found to hydrolyze the gelatin films most easily, while higher enzyme concentrations are required in case of pepsin, plasmin and collagenase. The exopeptidases, i.e. leucine aminopeptidase, amino acid arylamidase and carboxypeptidases A and B do not cause lysis of gelatin substrate films. The example of a rabbit blastocyst protease involved in implantation is given to demonstrate the application of gelatin substrate film tests for studies of enzymes which have no or little activity against known synthetic substrates (like BANA or GPNA) but hydrolyze gelatin films. Studies of interactions of this blastocyst protease with various inhibitors of known specificity, however, show that the active center of this enzyme nevertheless has striking similarities to trypsin (and also to chymotrypsin). The enzyme is possibly related to elastase. It is emphasized that, besides this, there are a number of different protease type enzymes in rabbit blastocyst and uterine tissues, some of which can be demonstrated only with chromogenic substrates and some only by gelatin methods. Aspects of applicability of the two types of protease tests are briefly discussed.
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PMID:[The specificity and sensitivity of the gelatin base protease substrate film test ]. 4 23

A cationic polypeptide growth factor, isolated from human serum and purified to homogeneity, has stimulated the replication of density-inhibited BALB/c 3T3 cells. It has a molecular weight of 1.3 x 10(4) daltons and an isoelectric point of 9.7. Trypsin or chymotrypsin digestion reduces the growth-stimulatory activity by 75%, whereas 2-mercaptoehanol completely abolishes it. The growth factor is heat-stable (100 degrees C X 10 min) and free of insulin-like activity. The highly purified serum growth factor has been labeled with 125I, and an antiserum to the growth factor was produced in the rabbit. A specific, highly sensitive radioimmunoassay has been developed. Factors with growth-stimulating activity have also been detected in human platelets and human pituitary gland extracts. Platelets and pituitary glands have antigenic determinants that are recognized by antibodies to the serum growth factor. The platelet and pituitary gland growth factors are also cationic and heat stable, and are destroyed by 2-mercaptoethanol. Thus the human serum, platelet, and pituitary gland growth factors have similar properties.
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PMID:Growth factors derived from human serum, platelets, and pituitary: properties and immunologic cross-reactivity. 8 59

Trypsin (T) and chymotrypsin (CHT) activities in luminal contents of the ileum, caecum and sigmoideum were followed in conventional (6 animals), monoassociated (5) and germfree (5) rabbits by pH-stat automatic titration using p-toluenesulphonyl-L-arginine methylester and acetyl-L-tyrosine ethylester as substrates. In conventional rabbits with complete microbial flora an aborally increasing decline of both proteolytic activities of luminal contents was determined (ileum T 198.2 - CHT 100.0; signmoideum T 10m.2 - CHT 68.8 mrg/g of intestinal content). Monoassociated animals represent a group different from both germfree and conventional animals. Trypsin and chymotrypsin of intestinal contents were not significantly altered by the presence of megacaecum in germfree rabbits (ileum T 219.2 - CHT 160.2; sigmoideum T 208.8 - CHT 110.8 mug/g of intestinal content). Chymotrypsin in the intestinal contents appears more labile and more affected by microbial flora than trypsin.
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PMID:Trypsin and chymotrypsin activity of the intestinal content in germfree, monoassociated and conventional rabbits. 13 38

Trypsin, thrombin, fibrinolysin, papain, chymothrypsin and urokinase were immobilized on aminopolystyrene resin by the reaction of diazocoupling. An activation of prothrombin and plasminogen and also hydrolysis of fibrin by immobilized enzymes were studied. The immobilized enzymes hydrolyzed N-benzoyl-1-arginine ethyl ester and L-tyrosine ethyl ester. The only preparation of immobilized thrombin possessed the coagulational activity. After the covalent binding trypsin and plasmin maintained the capacity to cause a fibrinolysis. Immobilized trypsin, plasmin, papain, chymotrypsin and urokinase exhibited the fibrinolytic effect due to convertion of plasminogen into plasmin.
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PMID:[Blood coagulating properties of immobilized proteases]. 14 May 25


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