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

---

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

Trypsin, porcine pancreatic kallikrein, and several blood coagulation enzymes, including bovine thrombin, bovine factor Xa, human factor Xa, human plasma factor XIa, human plasma factor XIIa, and human plasma kallikrein, were inactivated by a number of substituted isocoumarins containing basic functional groups (aminoalkoxy, guanidino, and isothiureidoalkoxy). 3-Alkoxy-4-chloro-7-guanidinoisocoumarins were found to be the most potent inhibitors for the coagulation enzymes tested with kobsd/[I] values in the range of 10(3)-10(5) M-1 s-1. 4-Chloro-3-isothiureidoalkoxyisocoumarins show high inhibitory potency toward porcine pancreatic kallikrein, human plasma kallikrein, human factor XIa, human factor XIIa, and trypsin with kobsd/[I] values of the order of 10(4)-10(5) M-1 s-1. The inhibition of these serine proteases by the substituted isocoumarins are time dependent, and the inactivation of trypsin by 3-alkoxy-4-chloro-7-guanidinoisocoumarins and 7-amino-4-chloro-3-(3-isothiureidopropoxy)isocoumarin occured concurrently with the loss of the isocoumarin absorbance. The complex formed from inactivation of trypsin by these two types of inhibitors was very stable and regained less than 4% activity in 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid buffer (pH 7.5) after 1 day at 25 degrees C and regained 8-45% activity upon addition of buffered 0.29 M hydroxylamine. Trypsin inactivated by other inhibitors regained full activity upon standing or addition of hydroxylamine. Thrombin inactivated by 3-alkoxy-4-chloro-7-guanidinoisocoumarins was also quite stable and only regained 9-15% activity under similar conditions. These results are consistent with a proposed mechanism, where serine proteases inactivated by aminoalkoxyisocoumarins or isothiureidoalkoxyisocoumarins form acyl enzymes that will deacylate upon standing or addition of hydroxylamine. However, the acyl enzymes formed from 3-alkoxy-4-chloro-7-guanidinoisocoumarins or 7-amino-4-chloro-3-(3-isothiureidopropoxy)-isocoumarin will decompose further, probably through a quinone imine methide, to give an irreversibly inactivated enzyme by reaction with an active-site nucleophile such as His-57. The quinone imine methide intermediate may also react with a solvent nucleophile to give an acyl enzyme that can be reactivated by hydroxylamine. The inhibitors 4-chloro-7-guanidino-3-methoxyisocoumarin and 4-chloro-3-ethoxy-7-guanidinoisocoumarin have been tested as anticoagulants in human plasma and were effective at prolonging the prothrombin time. However, they are unstable in plasma (t1/2 = 4-8 min), and their in vivo utility may be limited.
...
PMID:Mechanism-based isocoumarin inhibitors for trypsin and blood coagulation serine proteases: new anticoagulants. 316 17

A unique tissue kallikrein-binding protein was identified and partially characterized in the brain and serum of Sprague-Dawley rats and in the serum-free conditioned media of mouse anterior pituitary cells (AtT 20) and rodent neuroblastoma x glioma hybrids (NG108-15). Kallikrein and kallikrein-binding protein(s) form SDS- and heat-stable complexes with a molecular weight (Mr) of approximately 92,000. The complex formation of 125I-labelled kallikrein and the binding protein in the serum and brain is inhibited by excess unlabelled rat urinary kallikrein, rat arginine esterase A (a kallikrein-like kininogenase), and human urinary kallikrein. When the active site of kallikrein was blocked by phenylmethylsulfonyl fluoride or D-Phe-D-Phe-L-Arg-CH2Cl, no complex formation was detected. Kallikrein-binding protein only forms complexes with active kallikrein or trypsin-activated prokallikrein but not with prokallikrein. 125I-labelled kallikrein forms a 92-kilodalton protein with binding protein in various brain regions of perfused normotensive rats of the Wistar-Kyoto strain (WKY), including the cerebral cortex, cerebellum and brain stem; but complex formation was not found in corresponding brain regions of the spontaneously hypertensive rat (SHR). Similarly, the kallikrein-binding protein was identified in various tissues including thymus, lung, liver, prostate, Cowper's gland, adrenal gland, kidney, and pancreas of WKY rats but not in tissues of SHR. The results suggest a major difference in the kallikrein-binding protein in hypertensive versus normotensive rats. The role of this specific kallikrein-binding protein in cellular hemodynamic processes and blood pressure regulation remains to be investigated.
...
PMID:A major difference of kallikrein-binding protein in spontaneously hypertensive versus normotensive rats. 317 Nov 70

The measurement of glandular kallikrein in biological fluids most often utilizes a synthetic substrate, H-D-valylleucylarginine-p-nitroanilide (S-2266), which assesses amidase activity. Although this substrate has reasonable specificity for glandular kallikrein, other tryptic-like proteases found in mixed saliva may also cause hydrolysis. The primary purpose of this study was to assess the accuracy of the use of this substrate for the measurement of glandular kallikrein in human mixed saliva. An additional objective was to determine the presence of prekallikrein in mixed saliva. The addition of soybean trypsin inhibitor (SBTI), which inhibits other tryptic-like enzymes but not glandular kallikrein, resulted in an approx. 30 per cent decrease in the hydrolysis of S-2266 by centrifuged mixed human saliva. A correlation of 0.918 was obtained between the biological assays for kinin release and amidase activity in 19 subject samples. Amidase activity increased following treatment of saliva with trypsin, indicating the presence of prekallikrein in human mixed saliva. It is concluded that S-2266 is an accurate substrate for the assay of glandular kallikrein in human mixed saliva; that the inclusion of SBTI in the assay mixture is needed to inhibit non-kallikrein proteases that may also hydrolyse the synthetic substrate; and that prekallikrein is present in mixed saliva. Thus any future studies of changes in the level of kallikrein in saliva may wish to consider the presence of both active and total levels of glandular kallikrein.
...
PMID:The assay of glandular kallikrein and prekallikrein in human mixed saliva. 324 88

Inhibitors of trypsin and alpha-chymotrypsin with Mr of about 7000 Da and isoelectric points of greater than 10 and 9.9, respectively, were isolated from the venom of the common viper Vipera berus berus, using gel filtration and ion exchange chromatography. The inhibitor I prefers alpha-chymotrypsin (Ki = 4.6 X 10(-10) M) for the formation of an enzymeinhibitor complex at a molar ratio of 1:1. The inhibitor II prefers trypsin (Ki = 6.7 X 10(-11) M), forms an EI-complex at a molar ratio of 1:2, but also inhibits alpha-chymotrypsin (Ki = 1.4 X 10(-9) M) and hog pancreatic kallikrein (Ki = 1.6 X 10(-8) M). The inhibitor II contains no valine or methionine.
...
PMID:[Trypsin and chymotrypsin inhibitors from viper venom]. 325 36

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.
...
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.
...
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.
...
PMID:Identification of latent tissue kallikrein, prolactin and growth hormone secretion in GH3 pituitary cells using modified radioimmunoassays. 336 Feb 6

A protein-binding radioimmunoassay (RIA) of rat low molecular weight (LMW) kininogen with the following characteristics has been developed: sensitivity, 2.5 ng/tube; inter-assay coefficient of variation, 12.4% (N = 28); and intra-assay coefficient of variation, 9.4% (N = 11). The new assay correlated (r = 1) with the determination of kinin equivalence of kininogen after trypsinization. The cross-reactivity of rabbit anti-rat LMW kininogen antibody was 2.5% with bovine LMW kininogen, 5.8% with rat plasma high molecular weight (HMW) kininogen, and none with kinin. Although the antibody appears to partially recognize des-kinin-kininogen, the low degree of cross-reactivity and the extremely low levels of kinin-free-kininogen allow accurate determination of total LMW kininogen in rat urines. The LMW kininogen formed 20% kinins with salivary kallikrein when compared with trypsin, suggesting that the preparation consists of both K- and T-kininogens (K = kallikrein susceptible; T = trypsin susceptible). The newly developed protein-binding RIA recognizes LMW kininogen of rat urine which consists of both K- and T-kininogens.
...
PMID:A new direct radioimmunoassay of rat urinary kininogen. 337 4

Glandular kallikrein (a trypsin-like serine protease) is an estrogen-induced and dopamine-repressed protein in the rat anterior pituitary which appears to be associated with lactotrophs. This study examined glandular kallikrein levels in diethylstilbestrol (DES)-induced pituitary tumors in F344 rats and compared it to plasma and pituitary prolactin, and pituitary wet weight. Ovariectomized F344 rats were implanted with Silastic tubes containing 0 or 5 mg DES for 1, 3, 5, 7, or 9 weeks. Glandular kallikrein was measured by microenzymatic assay using D-valylleucylarginyl-p-nitroanilide following trypsin treatment of extracts to activate latent forms of glandular kallikrein. Prolactin was measured by radioimmunoassay. DES induced steady time-dependent increases in pituitary wet weight with 7- and 16-fold increases observed by 5 and 9 weeks, respectively. Growth rates averaged 11.4 mg/week during the first 5 weeks of DES exposure, and then increased to 23.2 mg/week between weeks 5 and 9. Glandular kallikrein total activity (nmol/min/pituitary) increased 130- and 240-fold after 3 and 5 weeks of DES exposure, respectively, and then abruptly plateaued. The specific activity (nmol/min/mg protein) of glandular kallikrein peaked at 3-5 weeks (36-fold increase compared to controls) and then declined as pituitary protein but not glandular kallikrein continued to increase. Total pituitary prolactin constantly rose during DES exposure with 12- and 26-fold increases after 5 and 9 weeks, respectively. Plasma prolactin levels also continuously rose during exposure to DES with 130- and 290-fold increases after 5 and 9 weeks, respectively. No major strain differences were found with regard to sensitivity to the acute effects of estrogen or dopaminergic stimulation on glandular kallikrein induction. DES-induced pituitary tumors in F344 rats are well known to arise via lactotroph proliferation, and the striking elevation in glandular kallikrein and prolactin during the early phases of tumor growth provide further support for a localization of glandular kallikrein in lactotrophs. However, the abrupt stabilization in glandular kallikrein levels by week 5 was unexpected and may signal a biochemical transformation of the tissue during tumor progression.
...
PMID:Glandular kallikrein in estrogen-induced pituitary tumors: time course of induction and correlation with prolactin. 339 Aug 8

The relationship between chemical modifications of arginine derivatives and inhibitory activity to horse serum cholinesterase (BuChE) was investigated. It provided a new insight into the topography of the active site of BuChE. 1) BuChE has the hydrophobic binding pocket, the depth of which corresponds to the length of ethylpiperidine. 2) In the opposite side to the hydrophobic binding pocket, BuChE has a certain entity which repulses carboxyl group at the 2-position of piperidine of L-arginine piperidine amide. 3) The P site of BuChE can allow 4-propyl and 4-phenyl group attached to piperidine. Comparison of the results with those of thrombin and trypsin clearly revealed similarities and dissimilarities among BuChE, trypsin, and thrombin in the active site topography, and hence, we introduce a new selective inhibitor for BuChE, N alpha-dansyl-L-arginine 4-phenylpiperidine amide. It inhibits BuChE strongly (Ki = 0.016 microM), whereas it inhibits trypsin, thrombin, plasmin, and glandular kallikrein only weakly and shows actually no inhibition on acetylcholinesterase from the human erythrocyte. In addition, the new inhibitor becomes highly fluorescent when bound with BuChE, indicating that the compound is an ideal probe of the interactions of BuChE as well as a titrant of it.
...
PMID:N alpha-dansyl-L-arginine 4-phenylpiperidine amide. A potent and selective inhibitor of horse serum cholinesterase. 340 26


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---