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)

The effect of tryptase, a neutral protease released from human lung mast cell secretory granules, on the tissue prokallikrein present in human urine was examined. Tryptase has been shown previously to lack activity against plasma prokallikrein. Purified tryptase was incubated with a concentrated preparation of urinary prokallikrein. No increase in kallikrein-like enzymatic activity or immunoreactive tissue kallikrein was detected. Activation of urinary prokallikrein with trypsin served as a positive control. Furthermore, preincubation of urinary prokallikrein with tryptase did not diminish the subsequent activation of urinary prokallikrein by trypsin. Therefore, tryptase neither activates nor destroys human tissue or plasma prokallikreins.
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PMID:Tryptase and kinin generation: tryptase from human mast cells does not activate human urinary prokallikrein. 329 74

Studies suggest that the actions of insulin on glucose metabolism may be mediated through activation of a membrane-bound serine protease with properties similar to a kallikrein-like enzyme. Also, bradykinin, a vasoactive product of kallikrein's action upon kininogen substrates, increases glucose uptake when infused into the human forearm. To determine whether a kallikrein or a kinin directly affects cellular glucose metabolism or participates in mediating insulin's actions, we studied their effects on isolated rat soleus muscle. Although trypsin (1.34 microM) increased incorporation of glucose into muscle glycogen to the same extent as insulin (200 mu units/ml), a purified rat tissue (urinary) kallikrein (0.4-1.34 microM) produced no such effect. Furthermore, the tissue kallikrein inhibitor, aprotinin, or a polyclonal kallikrein antiserum did not inhibit the action of insulin on incorporation of glucose into muscle glycogen. Treatment of the muscle preparation with bradykinin (1nM - 10 microM) did not result in any change in basal or insulin-stimulated (20 - 2000 mu units/ml) entry of glucose into glycogen or the glycolytic pathway. Bradykinin (1nM - 10 microM) also did not influence basal or insulin-stimulated (1000 mu units/ml) initial rates of glucose transport. These studies suggest that the previously observed in vivo effects of bradykinin on peripheral glucose uptake are probably mediated by changes in tissue perfusion rather than direct kinin effects on skeletal muscle, and that the putative membrane serine protease involved in the insulin-effector system is not tissue kallikrein.
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PMID:Tissue kallikrein and bradykinin do not have direct insulin-like actions on skeletal muscle glucose utilization. 332 22

Our studies demonstrate that rat anterior pituitary cells (GH3) are capable of synthesizing and secreting tissue kallikrein together with prolactin and growth hormone. The secretion of prolactin and growth hormone in GH3 cells was measured by two newly developed sensitive radioimmunoassays (RIA), using the polyethylene glycol separation technique. In the direct radioimmunoassay for rat tissue kallikrein, using a polyclonal antiserum which recognizes both active and prokallikrein, the GH3 kallikrein displays parallelism with standard curves of rat urinary kallikrein. The production of immunoreactive kallikrein, prolactin, and growth hormone is time-dependent, and the levels after a 72 h incubation in serum-free media are approximately 12.2 +/- 4.4 ng, 272.2 +/- 33.0 ng, and 475.6 +/- 4.8 ng per 10(6) cells per ml (mean +/- SD, n = 3), respectively. In Western blot analyses, a specific monoclonal antibody to tissue kallikrein (V4D11) identifies GH3-secreted kallikrein as a approximately 39,000 Da protein, slightly larger than approximately 38,000 Da kallikreins of submandibular gland, mouse anterior pituitary cells (AtT 20) or rodent neuroblastoma X glioma hybrid cells (NG108). Kallikrein mRNA in GH3 cells was identified in Northern blot analyses, using a tissue kallikrein cDNA probe. In a RIA using a kallikrein monoclonal antibody (V1C3) recognizing only active kallikrein, kallikrein could not be detected in the media incubated up to 48 h with GH3 cells. However, after trypsin treatment, a time-dependent increase of immunoreactive kallikrein (using monoclonal antibody V1C3), Tos-Arg-OMe esterase, and kinin-releasing activities can be measured in the conditioned media. The activated esterase activity was inhibited by aprotinin and by affinity-purified kallikrein monoclonal antibody (V4D11) in a dose-dependent manner. The data indicated that rat anterior pituitary GH3 cells secrete latent tissue kallikrein, which can be converted to active kallikrein by trypsin. These hormonally responsive cells co-synthesize kallikrein with prolactin and growth hormone and provide a model system for studying the regulation of kallikrein gene expression.
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PMID:Identification of latent tissue kallikrein, prolactin and growth hormone secretion in GH3 pituitary cells using modified radioimmunoassays. 336 Feb 6

Kininogen sequence analogs containing amino acid residues around the Arg-Ser cleavage site of bovine kininogens were prepared with bulky aliphatic residues in the P3 position as specific inhibitors of tissue kallikrein. KKI-7 (containing a cyclohexylacetyl group) and KKI-8 (containing an adamantaneacetyl group) both inhibited human urinary kallikrein with KI = 4 microM. These inhibitors are 40 times more potent than the corresponding peptide containing the naturally occurring prolyl residue at P3 and one-seventh as susceptible to hydrolysis. KKI-7 and KKI-8 are poor inhibitors of human plasma kallikrein (KI = 244 and 358 microM, respectively) while their capacity to inhibit trypsin is 1/3 and 1/17, respectively, that of their inhibitory capacity for human urinary kallikrein. When KKI-7 was tested in a rat blood flow model, the infused peptide (50 micrograms/kg/min) depressed the response to injected rat submandibular kallikrein (200-500 ng) to 52% of the control response when it was injected less than 10 minutes after infusion was started and to 16% of the control response when it was injected between 10 and 30 minutes after infusion of the inhibitor, the response gradually returned toward normal. Similar results were obtained with porcine pancreatic kallikrein. Infusion of the inhibitor did not affect the response to bradykinin and infusion of the vehicle itself did not alter the response to either injected kallikrein or bradykinin. These analogs have greater specificity and stability than those previously developed and are appropriate for the in vivo inhibition of glandular kallikreins.
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PMID:In vivo assay of specific kallikrein inhibitors. 348 Dec 13

Kinetic constants for the hydrolysis by porcine tissue beta-kallikrein B and by bovine trypsin of a number of peptides related to the sequence of kininogen (also one containing a P2 glycine residue instead of phenylalanine) and of a series of corresponding arginyl peptide esters with various apolar P2 residues have been determined under strictly comparative conditions. kcat and kcat/Km values for the hydrolysis of the Arg-Ser bonds of the peptides by trypsin are conspicuously high. kcat for the best of the peptide substrates, Ac-Phe-Arg-Ser-Val-NH2, even reaches kcat for the corresponding methyl ester, indicating rate-limiting deacylation also in the hydrolysis of a peptide bond by this enzyme. kcat/Km for the hydrolysis of the peptide esters with different nonpolar L-amino acids in P2 is remarkably constant (range 1.7), as it is for the pair of the above pentapeptides with P2 glycine or phenylalanine. kcat for the ester substrates varies fivefold, however, being greatest for the P2 glycine compounds. Obviously, an increased potential of a P2 residue for interactions with the enzyme lowers the rate of deacylation. In contrast to results obtained with chymotrypsin and pancreatic elastase, trypsin is well able to tolerate a P3 proline residue. In the hydrolysis of peptide esters, tissue kallikrein is definitely superior to trypsin. Conversely, peptide bonds are hydrolyzed less efficiently by tissue kallikrein and the acylation reaction is rate-limiting. The influence of the length of peptide substrates is similar in both enzymes and indicates an extension of the substrate recognition site from subsite S3 to at least S'3 of tissue kallikrein and the importance of a hydrogen bond between the P3 carbonyl group and Gly-216 of the enzymes. Tissue kallikrein also tolerates a P3 proline residue well. In sharp contrast to the behaviour of trypsin is the very strong influence of the P2 residue in tissue-kallikrein-catalyzed reactions. kcat/Km varies 75-fold in the series of the dipeptide esters with nonpolar L-amino acid residues in P2, a P2 glycine residue furnishing the worst and phenylalanine the best substrate, whereas this exchange in the pentapeptides changes kcat/Km as much as 730-fold. This behaviour, together with the high value of kcat/Km for Ac-Phe-Arg-OMe of 3.75 X 10(7) M-1 s-1, suggests rate-limiting binding (k1) in the hydrolysis of the best ester substrates.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Effects of secondary interactions on the kinetics of peptide and peptide ester hydrolysis by tissue kallikrein and trypsin. 364 48

A series of acetyl-peptidyl-amides containing the amino acid sequence around the Arg-Ser kallikrein cleavage site of bovine kininogen were synthesized and tested for their ability to inhibit both the kinin-releasing activity and the amidase activity of purified human urinary kallikrein. The substrate analogues were competitive inhibitors for human urinary kallikrein and the heptapeptides (P4-P3'), hexapeptides (P3-P3'), and pentapeptides (P2-P3') gave Ki values of 140, 64, and 18 microM respectively, while the tetrapeptides (P1-P3'), tripeptides (P1'-P3') and dipeptides (P2'-P3') had little or no inhibitory activity. The effective analogues had neither kinin-like nor kinin-blocking activity on the rat uterus either before or after exposure to human urinary kallikrein. The effective human urinary kallikrein inhibitors were further examined for their effect on other serine proteases, including human plasma kallikrein, plasmin, complement components (C1s, C1r), bovine coagulation factors (IIa, IXa, and Xa), elastase, and trypsin. These peptides showed little inhibition of the circulating serine proteases but yielded a Ki for the nonspecific protease trypsin in the microM range. These results should provide the basis for the development of highly specific tissue kallikrein inhibitors to aid in elucidating the in vivo role(s) of tissue kallikreins.
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PMID:Specificity of substrate analogue inhibitors of human urinary kallikrein. 384 67

Based on a study of the kininogenase activity of the total plasma kallikrein in the presence of 3 concentrations of the soybean inhibitor trypsin (0.5, 1.0, 10.0 micrograms/ml) one can measure at a time the activity of tissue kallikrein (without specifying the source) and the activity of 3 forms of plasma kallikrein, including its adsorption on kaolin that characterizes the conformational structure of the enzyme. Examination of 10 healthy subjects and 136 patients revealed a 10 to 20-fold increase in the content of tissue kallikrein in plasma of 70% of diabetes mellitus patients and a 2.5 to 3-fold elevation in 50% of patients with chronic occupational bronchitis, and in 30% of patients suffering from chronic hepatitis. The method suggested makes it possible to have a better insight into the physiological and pathogenetic role of the kinin system and may be used for laboratory control over the treatment efficacy.
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PMID:[Method for determining kallikrein of tissue origin in blood plasma and its clinical significance]. 384 14

The rate of tissue kallikrein (EC 3.4.21.35) excretion into the urine has been examined with an active site-specific radioimmunoassay for kallikrein in renal transplant recipients, in post-uninephrectomy kidney donors, and in a normal control population. Normal individuals on uncontrolled diets excreted 96.88 +/- 7.00 (SEM) micrograms of active kallikrein/24 hr and 113.68 +/- 8.39 micrograms of total kallikrein/24 hr, as determined after trypsin treatment of urine samples. Uninephrectomized donors secreted significantly less (P less than 0.05) active (44.99 +/- 6.39 micrograms/24 hr) and total (73.59 +/- 11.95 micrograms/24 hr) kallikrein than either the entire normal population or an age-matched subpopulation. Recipients with good renal function who had received kidneys 2 to 13 years prior to kallikrein assay excreted less (P less than 0.05) active (13.21 +/- 2.50 micrograms/24 hr) and total (18.69 +/- 3.65 micrograms/24 hr) kallikrein than either normal or uninephrectomized populations. Similar values for active (11.05 +/- 1.56 micrograms/24 hr) and total (17.60 +/- 1.96 micrograms/24 hr) kallikrein were seen in patients who had received kidneys within 6 months of assay. Thus, kallikrein excretion in kidney recipients remains significantly lower than in uninephrectomized donors. As compared to normal individuals, the reduced kallikrein excretion in post-uninephrectomized kidney donors and in renal allograft recipients suggests that renal kallikrein excretion may reflect functional distal tubular mass.
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PMID:Kallikrein excretion in renal transplant recipients and in uninephrectomized donors. 390 May 31

Polyvalent peptide-protein inhibitors of proteinases trasilol, contrical, hordox and pantrypine were studied using purified preparations of kallikreins from human blood plasma and hog pancreas as well as of trypsin. All the trade grade preparations of the inhibitors studied exhibited more distinct affinity to trypsin and tissue kallikrein as compared with blood plasma kallikrein. Trasilol inhibited most effectively the activity of trypsin, contrical--of pancreatic kallikrein. Contrical, trasilol and hordox inactivated blood plasma kallikrein at similar rates, pantrypine was several times less effective.
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PMID:[Effectiveness of commercial polyvalent proteinase inhibitors. Comparative analysis of their effect on kallikreins from different sources and trypsin]. 616 84

Fab fragments from two new monospecific anti-human tissue kallikrein sera were examined for their capacity to inhibit the functional activities of purified human urinary kallikrein and purified human pancreatic kallikrein. Fragments from a new anti-urinary kallikrein serum and from an anti-pancreatic kallikrein serum yielded mixed inhibition of kinin-generating activity and minimal inhibition of esterolytic activity. In contrast to the previously described "active site directed" anti-urinary kallikrein, these new antisera demonstrated little specificity for epitopes near the enzymatic site of urinary or pancreatic kallikrein. When used to localize kallikrein antigen in human pancreas obtained at surgery, IgG fractions of the new anti-kallikrein sera yielded moderate acinar and ductal staining in the absence of pretreatment of the tissue with trypsin or pronase. Short incubation with 0.125 mg/ml of either enzyme permitted the discrete localization of islet beta cell kallikrein antigen, while increased pronase concentrations decreased kallikrein antigen in both islets and exocrine tissue and led to islet destruction. Both antibody specificity and tissue preparation influence kallikrein localization in human pancreas.
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PMID:Immunohistochemical localization of glandular kallikrein in the endocrine and exocrine human pancreas. 619 4


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