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:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Anhydrotrypsin, a derivative of bovine trypsin, immobilized on Sepharose tightly adsorbs various peptides containing L-arginine at the carboxyl termini, such as bradykinin and tuftsin. These peptides correspond to the specific products of the action of trypsin-like enzymes. Native trypsin immobilized on Sepharose does not show such strong affinity. Fragment 2, a peptide with 41 amino acid residues, which has been released together with bradykinin from bovine high-molecular-weight kininogen by the action of plasm kallikrein, is also adsorbed on the immobilized anhydrotrypsin. When only the carboxyl-terminal arginine is removed with carboxy-peptidase B, however, the peptide loses its adsorptive ability. Immobilized anhydrochymotrypsin, on the other hand, exerts specific affinity for the peptides which correspond to the products of chymotrypsin. These results suggest that the anhydro-derivatives of serine-proteseas in general may be of great use in the affinity chromatography of respective series of various naturally occurring peptides.
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PMID:Specific isolation of biologically-active peptides by means of immobilized anhydrotrypsin and anhydrochymotrypsin. 57 60

Human mucous secretions contain low molecular weight (Mr approximately 11,000) acid-stable inhibitors directed against elastase and cathepsin G from PMN-granulocytes. Important biochemical properties of these inhibitors are presented and their possible biological function is discussed. An inhibitor of glandular and plasma kallikreins preventing kinin-liberation from kininogen but not ester hydrolysis was obtained from rat kidney tubules. A molecular weight of about 4700 was estimated for this kallikrein-specific inhibitor (trypsin-induced kinin-liberation is not prevented).
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PMID:Naturally occurring low molecular weight inhibitors of neutral proteinases from PMN-granulocytes and of kallikreins. 63 56

Human trypsins 1 and 2 both converted Met-Lys-bradykinin to bradykinin and released bradykinin from kininogen in human plasma as measured by bioassay with the isolated guinea pig ileum. Porcine kallikrein did not act on Met-Lys-bradykinin and released kallidin from human kininogen. Since human trypsin 1 is only partially and trypsin 2 completely inhibited by soybean trypsin inhibitor, these data show that the criterion of susceptibility to soybean trypsin inhibitor cannot be used to discriminate between trypsin and kallikrein of different species.
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PMID:Comparison of the kininogenase activity of human pancreatic trypsins and porcine Kallikrein on Met-Lys-bradykinin and human plasma kininogen. 71 Nov 61

Synthetic procedures have been developed for the preparation of peptides of arginine chloromethyl ketone and applied in the preparation of affinity labels which correspond to the -Pro-Phe-Arg- C terminus of bradykinin, a physiological cleavage site of kallikrein in kininogen. Two such reagents, Ala-Phe-ArgCH2C1 and Pro-Phe-ArgCH2C1, proved to be highly effective as well as selective affinity labels for human plasma kallikrein. For example, Pro-Phe-ArgCH2C1 inactivates plasma kallikrein 50% in 24 min at a concentration of 2 x 10(-8)M, while other trypsin-like proteases are less susceptible in inactivation than kallikrein, differing by a factor of 48 for plasmin and factors of 10(2)-10(5) for factor Xa, thrombin, and urokinase. The affinity of human plasma kallikrein for Ala-Phe-ArgCH2C1 (Ki = 0.078 micron) is about 60 times that for Ala-Phe-LysCH2C1(Ki = 4.9 micron), whereas human plasmin exhibits about the same affinity for the former affinity label (Ki = 1.3 micron) as for the latter (Ki = 0.83 micron). The rate constants for the irreversible step of the affinity labeling reaction, k2, are similar for affinity labels tested with the individual proteases: 0.35 min-1 for plasma kallikrein and 0.18 min-1 for plasmin.
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PMID:Synthesis of peptides of arginine chloromethyl ketone. Selective inactivation of human plasma kallikrein. 72 86

Recent studies of individuals with high molecular weight (HMW) kininogen deficiency established the importance of this plasma protein for in vitro initiation of blood coagulation. In the present study, HMW-kininogen was highly purified from human plasma by monitoring its clot-promoting activity, using Fitzgerald trait plasma as a substrate. This preparation of HMW-kininogen revealed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (mol wt: 120,000) and released 1% of its weight as bradykinin upon incubation with plasma kallikrein. HMW-kininogen specifically repaired impaired surface-mediated plasma reactions of Fitzgerald trait plasma, but did not affect those of Hageman trait and Fletcher trait plasma. Kinin release from HMW-kininogen by trypsin, but not by plasma kallikrein, resulted in total loss of clot-promoting activity. No inhibitors of coagulation were found when all kinin activity was removed from HMW-kininogen by trypsin. The roles of HMW-kininogen, Hageman factor (HF, Factor XII), plasma prekallikrein (Fletcher factor), and plasma thromboplastin antecedent (PTA, Factor XI) in blood coagulation were studied in a purified system. HMW-kininogen was absolutely required for activation of PTA by HF and ellagic acid. The yield of activated PTA was proportional to the amount of HF, HMW-kininogen, and PTA in the mixtures, suggesting that, to activate PTA, these three proteins might form a complex in the presence of ellagic acid. No fragmentation of HF was found under these conditions. In contrast to HF, HF-fragments (mol wt: 30,000) activated PTA in the absence of HMW-kininogen and ellagic acid. Thus, it appears that in the present study PTA was activated in two distinct ways. Which pathway is the major one in whole plasma remains to be determined.
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PMID:Purification of high molecular weight kininogen and the role of this agent in blood coagulation. 89 64

The kinetic constants for horse urinary kallikrein and trypsin hydrolysis of BAEE, TAME, bradykinin methyl ester and bradykinyl-Ser-Val-Gin-Val-Ser were determined. The values of the ratio kcat/Km show that (1) kallikrein is catalytically less efficient than trypsin for all the substrates (2) the three esters are equally good substrates for trypsin while horse urinary kallikrein is 100-fold more effective on bradykinin methyl ester than on the other substrates (3) for both enzymes the ester of bradykinin is a better substrate than the tetradecapeptide.
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PMID:Kinetics of the hydrolysis of synthetic substrates by horse urinary kallikrein and trypsin. 98 54

Plasma kininogen, a labile precursor of bradykinin long considered difficult to characterize, has been assayed and partially purified by accelerated, non-destructive methods. Although heparin minimized premature kininogen consumption better than hexadimethrine bromide, both were ineffective when used in combination. The effects of varying incubation parameters upon kininogen consumption by trypsin were studied. Since trypsin (50 mug/ml) liberated in 10 min at 45degreesC in 0.1 M CaC12 as much bradykinin as in 30 min at 37 degrees C, the former conditions were adopted for assaying kininogen in terms of bradykinin equivalents released, as determined by rat uterus bioassay. The fastest (2-day) and simplest procedure for a routine 10-fold purification of kininogen from plasma consisted of ammonium sulfate precipitation (33-46%) followed by a single passage through a concanavalin A-agarose column. Con A binds glycoproteins (e.g. kininogen) more firmly than other proteins (albumin, gamma-globulins). Kininogen, resistant to killikrein attack while bound, was desorbed with 0.05 M alpha-methyl mannoside.
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PMID:Accelerated assay, preservation and purification (concanavalin A) of plasma kininogen. 99 66

A preparation of high-molecular weight kininogen (HMWK) was isolated from rabbit citrate blood plasma and purified using chromatography on DEAE-Sephadex A-50 and CM-Sephadex C-50. From 1 mg HMWK, trypsin or kallikreine of human blood plasma release 10 mkg bradykinine. The HMWK preparation is homogeneous during electrophoresis in 7.5% polyacrylamide gel in tris-glycine buffer, pH 8.3; its electrophoretic mobility corresponds to that of alpha2-globulins. The molecular weight of HMWK estimated using the collumn with Sephadex G-200, is 130.000--150.000; the sedimentation constant S20w is 7.6. Rabbit HMWK is neither a dimer, nor a trimer of low molecular weight kininogen (LMWK), since it does not degrade into subunits after treatment by 2.5% solution of sodium dodecyl sulfate, containing 8 M urea. 0.05 M 2-mercaptoethanol and 8 M urea induce HMWK splitting into 2 fragments with respective molecular weights of 80.000 and 30.000, the kinine-containing group being localized in the low-molecular weight fragment. Estimation of rates of kinine formation by different kininogenasses from highly purified HMWK and LMWK preparations showed that those kininogens are functionally different substrates, since blood plasma kallikreines release kinines from HMWK at a greater rates, whereas tissue kallikreines, e. g. human saliva kallikreine release kinines from LMWK. The specificity of kallikreines as kininogenase, to trypsin, was determined. Tripsin removes bradykinine from both kininogens at the same rates, which are an order of magnitude less than those found for kallikreines.
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PMID:[Purification and properties of high-molecular-weight rabbit kininogen]. 102 87

Plasma supernatant in which kallikrein has been activated and removed by glass powder whilst kininogen I (HMW) has been consumed by the activated kallikrein, was used for the preparation of kininogen II. It was purified by chromatography on DEAE-cellulose followed by gel filtration on G-200 Sephadex. The purification of kininogen II was assessed from determinations of the amount of kinin released (expressed as bradykinin) as measured on the isolated guinea pig ileum, using samples incubated with human salivary kallikrein or trypsin. A preparation of kininogen II containing an activity equivalent to 8 microgram Br/mg protein, was obtained. Salivary kallikrein released approximately three times more kinin from the substrate as compared to trypsin.
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PMID:[Purification of kininogen II (LMW) from human plasma]. 103 67

Kininogens I (HMW) and II (LMW) were isolated and partially purified from human plasma. The disappearance of kininogen I was prevented by the use of hexadimethrine bromide, which inhibits the activation of plasma prekallikrein. The two kininogens were separated by chromatography on DEAE-cellulose. Further purification of kininogen I and II was achieved by separate chromatographic steps of the partially purified kininogens on SP-Sephadex. The purification of the kininogens was controlled by incubation of the respective samples with different kininogenases: human plasma kallikrein, human salivary kallikrein and trypsin. As determined by gel filtration, a molecular weight of 250 000 daltons was found for kininogen I (HMW) and 48 000 daltons for kininogen II (LMW).
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PMID:[Purification and enzyme study of kininogens I and II from human plasma]. 103 68


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