Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.86 (clotting enzyme)
 176 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The milk-clotting enzyme pepsin was immobilized onto beads of alumina, titania, glass, stainless steel, iron oxide, and Teflon for treating skim milk in a fluidized-bed reactor. Two covalent attachment procedures using silanized supports and glutaraldehyde and two adsorption procedures were evaluated. The three best catalysts were titania and glass, using the covalent attachment procedure, and alumina, using the adsorption procedure at pH 1.2. The pepsin adsorbed on alumina catalyst has commercial potential compared to the previously used glass catalyst. Attempts to increase the stability of pepsin adsorbed on alumina by cross-linking with glutaraldehyde were unsuccessful owing to the low pH necessary for optimum pepsin adsorption; Desorption of pepsin from alumina during reactor operation was determined. Regeneration of spent catalysts was only partially successful.
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PMID:Pepsin immobilized on inorganic supports for the continuous coagulation of skim milk. 1 78

An active milk-clotting enzyme was purified some 40-fold from culture supernatant of Aspergillus ochraceus. The purification steps included ammonium sulfate precipitation, G-100 Sephadex gel filtration, and ion exchange chromatography, using DEAE Cellulose column. The enzyme exhibited milk-clotting activity and proteolytic behaviour, an optimum at pH 6.0 and in the range of 7--8.5, respectively. The purified enzyme was actively proteolytic against casein, haemoglobin, and bovine serum albumin at pH 8. The milk-clotting activity was greatly enhanced by manganous ions and by increasing concentrations of calcium chloride. Copper, zinc, and ammonium ions were potent inhibitors of the milk-curdling activity of the purified enzyme. Significant inhibition was also noted with sodium chloride at concentrations of 3% or more. Under the specified reaction condition, maximum rate of proteolysis against casein was obtained at 0.4% substrate concentration, whereas the milk-clotting time was linear proportional to dry skim milk concentration in the range of 8 to 24%. The results are discussed in comparison with other microbial milk-clotting enzymes, and limitations of applicability are also presented.
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PMID:Purification and properties of rennin-like enzyme from Aspergillus ochraceus. 3 45

The production of a milk-clotting enzyme by Aspergillus versicolor in 19 different culture media was investigated. Considerable milk-clotting activity was achieved by supplying corn steep liquor with either glucose of maltose. Dephytinization of corn steep liquor had an adverse effect on the production of milk-clotting enzyme. The results indicated that complex organic compounds favoured the production of the enzyme. Precipitation with acetone or tannin was unsuitable, but ammonium sulphate and ethanol above certain concentration produced active fractions.
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PMID:Production and isolation of milk-clotting enzyme from Aspergillus versicolor. 12 98

A pro-clotting enzyme capable of causing the gelation of clottable proteins in Limulus polyphemus (horseshoe crab) has been purified to apparent homogeneity as judged by sodium dodecyl sulfate-gel electrophoresis. The activation of the pro-clotting enzyme depended on the presence of both Ca+ and endotoxin. It contained gamma-carboxyglutamic acids and gave a single NH2-terminal lysine. The enzyme was inhibited by diisopropyl fluorophosphate, phenylmethylsulfonyl fluoride, and soy bean trypsin inhibitor, indicating that it is a serine protease. The molecular weight of the proclotting enzyme was determined to be at least 150,000 by sodium dodecyl sulfate-gel electrophoresis under reducing and denaturing conditions. The protein appears to consist of a single peptide chain, since exposure of the reduced and carboxymethylated enzyme to 6 M guanidine hydrochloride failed to dissociate it into any subunits.
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PMID:Studies on Limulus amoebocyte lysate. Isolation of pro-clotting enzyme. 32 51

Six commercial milk clotting preparations from animal and fungal sources were used to make cheddar cheese. The cheeses were analyzed initially and over 6-mo ripening for proximate composition, minerals, amino acids, soluble protein, nonprotein nitrogen, free fatty acids, lactones, and flavor development. No significant differences in the composition of the cheeses could be attributed to the type of clotting enzyme. One lot of one enzyme showed increased lipolytic activity which indicated contamination and suggested that the purity of the enzyme preparation should be checked.
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PMID:Composition of cheddar cheese made with different milk clotting enzymes. 33 10

A coagulogen in Limulus lysate has been purified to apparent homogeneity as judged by sodium dodecyl sulfate-gel electrophoresis. The purified coagulogen identified by its ability to clot with either trypsin or the purified Limulus clotting enzyme (Tai, J.Y., and Liu, T. Y. (1976) Fed. Proc. 35, 1486) has a molecular weight of 24,500. It consists of a single polypeptide chain of about 220 amino acids with glycine and serine as its NH2- and COOH-terminal residues; respectively. When acted upon by the Limulus clotting enzyme, the coagulogen releases a soluble C-peptide of about 45 amino acids and an insoluble coagulin of about 170 amino acids. The latter interacts in a noncovalent fashion to form the clot. Amino acid analyses together with the results of NH2- and COOH-terminal analyses suggest that the clot formation involves the cleavage of an--Arg--Lys--bond. Trypsin acts on the coagulogen to cause clotting by splitting the same peptide bond.
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PMID:Studies on Limulus amoebocyte lysate II. Purification of the coagulogen and the mechanism of clotting. 87 16

Bovine prothrombin was activated, in both the absence and presence of dissopropyphosphofluoridate (DEP) and benzamidine, by an activator which was highly purified from the venom of Echis carinatus (saw-scaled viper, ECV). The process of activation was monitored by sodium dodecysulfate (SDS)-polyacrylamide gel electrophoresis, and the reaction products were isolated and chemically characterized. In the absence of the inhibitors, prothrombin yielded two fragments with molecular weights of 28,000 and 57,000, of which the former was the N-terminal fragment of the zymogen and the latter was intermediate 1, consisting of a single polypeptide chain. Intermediate 1 was subsequently converted to an active intermediate, named intermediate ECV, without decrease of molecular weight. This new intermediate ECV, which showed little clotting activity but a strong alpha-N-tosyl-L-arginine methyl ester (TAME)-esterolytic activity and which bound with hirudin or antithrombin III, consisted of two polypeptide chains with molecular weights of 35,000 of 27,000 daltons. The former was indentified as the thrombin B chain with the N-terminal sequence Ile-Val-Glu-Gly and C-terminal serine, and the latter was a fragment with N-terminal Ser-Gly-Gly, linked to the thrombin A chain. On prolonged incubation, intermediate ECV autocaralytically yielded a fragment (inner fragment) of 14,000 daltons with N-terminal serine and the clotting enzyme alpha-thrombin [EC 3.4.21.5], which consists of A and B chains. In the presence of the inhibitors, intermediate ECV and the N-terminal fragment were accumulated in the activation mixture. On the other hand, when prothrombin was activated by the venom activator in the presence of hirudin, antithrombin III, or p-nitrophenyl p'-guanidinobenzoate, it did not yield any fragments but was converted to a derivative with two polypeptide chains having molecular weights of 51,000 and 34,000 daltons, of which the former consisted of N-terminal fragment, the inner fragment, and thrombin A chain, and the latter was thrombin B chain. This new prothrombin derivative, named prothrombin ECV, formed a high-molecular-weight complex, associating with antithrombin III. The complex was not dissociable even in the presence of SDS. Moreover, prothrombin ECV reacted with p-nitrophenyl p'-guanidinobenzoate. On the basis of the results described above, the mechanism of activaton of prothrombin by Echis carinatus venom activator can be summarized as follows: The venom activator first cleaves an Arg-Ile bond liniking thrombin A and B chains in the zymogen molecule, forming an active derivative, prothrombin ECV. This active derivative converts autocatalytically to intermediate ECV, liberating the N-terminal fragment, and active intermediate ECV generates alpha-thrombin, releasing the inner fragment. Thus, only a single peptide bond cleavage along the polypeptide chain of prothrombin is associated with activation by the venom activator...
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PMID:The mechanism of activation of bovine prothrombin by an activator isolated from Echis carinatus venon and characterization of the new active intermediates. 95 36

A clottable protein (coagulogen) isolated from a hemocyte lysate of the Japanese horseshoe crab (Tachypleus tridentatus) was incubated with an endotoxin-activated clotting enzyme(s) partially purified from the same lysate, and its structural change during gel formation was examined. The results indicated that the enzymatic formation of gel involved limited proteolysis of the Arg-Gly and Arg-Thr linkages located in the N-terminal portion of the coagulogen, liberating peptide C.
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PMID:A clottable protein (coagulogen) of horseshoe crab hemocytes. Structural change of its polypeptide chain during gel formation. 97 58

The effect of the aromatic diamidine derivative 2,6-bis (4-amidinobenzyl)-cyclohexanon-(1) on blood coagulation and fibrinolysis in vitro and in vivo was compared with that of the benzamidine derivative 4-amidinophenyl pyruvic acid and the aromatic diamidine derivative 4,4'-diamidinophenoxypentane. 2,6-Bis(4-amidinobenzyl)-cyclohexanon-(1) was found to be a strong inhibitor of the clotting enzyme thrombin. Because of the toxic side effects and pharmacokinetic properties of both diamidine derivatives their in vivo use as anticoagulants is limited.
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PMID:[Effects of aromatic bisamidines on blood coagulation and fibrinolysis]. 98 14

A turbidimetric assay for clottable plasma fibrinogen which is not sensitive to heparin or Pyran inhibition is described. The basis of the assay is the substitution of Reptilase-R for the thrombin usually employed. The assay correlates very well with a thrombin turbidimetric method and also has other advantages, including better stability of the clotting enzyme and more rapid attainment of equilibrium.
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PMID:A rapid, quantitative determination of clottable fibrinogen unaffected by heparin. 111 33


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