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:3.4.21.1 (chymotrypsin)
10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The purpose of this study was to assess the relationship of neuropeptide nerves and inflammatory leukocytes in PVG rats with adjuvant-induced arthritis. Substance P- and calcitonin gene-related peptide (CGRP)-immunoreactive nerves and inflammatory leukocytes were studied, using peroxidase (ABC) and/or alkaline phosphatase (APAAP) staining. Inflamed synovial tissue proper was infiltrated with neutrophils, ED1 macrophages and focal accumulations of CD2 T lymphocytes. In such tissue, the relationship between peptide-immunoreactive nerves and inflammatory cells was such that substance P and CGRP nerves were absent in heavily infiltrated villous synovial tissue, whereas healthy synovial tissue and non-inflammatory areas in adjuvant arthritic rats were innervated by substance P and CGRP nerves close to normal synovial tissue resident cells. In order to elucidate an eventual mechanism for lost immunoreactivity, healthy synovial tissue was exposed to chymotrypsin or oxygen derived free radicals (ODFR) in vitro. The former treatment caused total loss of immunoreactivity. These findings suggest that neuropeptides and neuropeptide containing nerves may be destroyed by locally produced proteolytic enzymes and various reactive oxygen species in the vicinity of inflammatory cells.
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PMID:Relationship between neuropeptide immunoreactive nerves and inflammatory cells in adjuvant arthritic rats. 137 4

Parathyroid hormone (PTH) -degrading activity was studied using osteoblast-like UMR-106 cells. PTH-degrading activity was assessed by the amount of PTH fragments produced in the medium after exposure of intact human PTH-(1-84) to UMR-106 cells. PTH immunoreactivity recovered in trichloroacetic acid-soluble products of the medium and in fractions eluted from reverse-phase high-performance liquid chromatography (HPLC) was measured by radioimmunoassay using an antibody specific for the mid-region and C-terminus of PTH. In this study, intact UMR-106 cells but not extracellular enzymes cleaved human PTH(1-84) into fragments which were released into the medium (in a time- and temperature-dependent fashion). HPLC analysis of the PTH fragments depicted three immunoreactive peaks (peaks 1, 2 and 3) besides intact PTH, indicating a limited PTH-hydrolyzing activity of the cells. Furthermore, a 1000-fold molar excess of either hPTH-(3-34) or [Nle8,Nle18,Tyr34]hPTH-(3-34)amide inhibited PTH-degrading activity by 63% and 80% of control, respectively, whereas neither calcitonin, vasopressin nor growth hormone suppressed it. Additionally, HPLC analysis of the samples treated with [Nle8,Nle18,Tyr34]hPTH-(3-34)amide showed a reduction of the three peaks, suggesting an involvement of PTH receptor in the production of PTH fragments. This PTH-degrading activity was strongly inhibited by phenylmethylsulfonyl fluoride and chymostatin, but not by soybean trypsin inhibitor, elastatinal or inhibitors of cysteine, aspartic or metalloproteinases, indicating that it is due to a seryl chymotrypsin-like endopeptidase. Chymotrypsin-like activity seems to be solely responsible for PTH-degrading activity in intact UMR-106 cells, since all three PTH fragments were predominantly suppressed in the presence of chymostatin. Further analysis of chymotrypsin-digested products of hPTH-(1-84) eluted from HPLC exhibited five fragments detected by ultraviolet absorbance at 210 nm, three of which were measurable by PTH radioimmunoassay, each corresponding to the three PTH fragments produced by UMR-106 cells. To explore the cleavage sites of PTH further, amino acid analysis of chymotrypsin-cleaved products was performed. The results strongly support the view that the chymotrypsin-like enzyme in UMR-106 cells cleaved the hormone between residues 23-24 and 34-35, to produce, at least, hPTH-(24-84) and -(35-84). Our present study indicates that a chymotrypsin-like endopeptidase is solely responsible for limited hydrolysis of PTH by intact UMR-106 cells.
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PMID:Parathyroid hormone degradation by chymotrypsin-like endopeptidase in the clonal osteogenic UMR-106 cell. 291 1

The effects of intra- and extraluminal capsaicin administration were evaluated in isolated perfused rat mesenteric bed. Capsaicin (10 nM-1 microM) produced a potent concentration-dependent relaxation of the tonic vasoconstriction induced by norepinephrine (1 microM) but not by high-K+ (60 mM). The capsaicin-induced relaxation was nearly abolished in preparations pretreated in vitro with a high concentration of capsaicin (1 microM, for 10 min, 1 h before). Capsaicin-induced relaxation was reduced but not abolished in preparations obtained from rats pretreated neonatally with capsaicin. The capsaicin effects were unaffected by atropine, guanethidine, propranolol, hexamethonium or tetrodotoxin. The observation that capsaicin (0.1 microM)-induced relaxation was virtually abolished in presence of the proteolytic enzyme alpha-chymotrypsin (1 U/ml) supports the involvement of neuropeptide(s) in this response. Bolus injections of calcitonin gene-related peptide (CGRP) elicited a potent and rapidly ensuing relaxation which underwent tachyphylaxis. However, no cross-desensitization with capsaicin was observed. It is concluded that activation of capsaicin-sensitive sensory fibers could release neuropeptides locally with a potent effect on intestinal blood flow.
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PMID:Vascular effects of capsaicin in isolated perfused rat mesenteric bed. 337 70

Messenger RNA extracted from rat medullary carcinoma of the thyroid directs the synthesis in cell-free translation systems of a precursor of calcitonin, Mr = 15,000, substantially larger than the mature form of the hormone, Mr = 3,500. When translations of the mRNA were carried out in the presence of microsomal membranes prepared from a canine pancreas, a larger product (apparent Mr = 17,000) was observed by electrophoresis of the labeled proteins in the translation mixtures on sodium dodecyl sulfate-polyacrylamide gels. This membrane-processed product of Mr = 17,000 was specifically immunoprecipitated by an antiserum to synthetic calcitonin and bound to concanavalin A-Sepharose. Incubation of the proteins synthesized in the cell-free translations performed in the presence of microsomal membranes with the glycosidase, endo-beta-N-acetylglucosaminidase H, reduced the apparent molecular weight of the membrane-processed precursor from 17,000 to 12,000. In addition, the processed Mr = 17,000 calcitonin-related precursor, but not the initial, unprocessed precursor of Mr = 15,000, was resistant to proteolytic digestion by a mixture of trypsin and chymotrypsin. These results indicate that the biosynthesis of calcitonin involves the glycosylation and proteolytic cleavage of a newly synthesized precursor along with sequestration of the processed precursor within microsomal vesicles. Thus, the calcitonin precursor undergoes extensive co- and post-translational processing to the smaller, unglycosylated hormone that is secreted.
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PMID:Procalcitonin is a glycoprotein. 720 75

Mediation of postprandial pancreatic enzyme secretion has been ascribed mainly to cholecystokinin and to vagovagal reflexes. Recent studies suggest that these pathways are subject to feedback regulation. Diversion of pancreatic juice from the duodenum stimulates cholecystokinin release and pancreatic enzyme secretion, and intraduodenal administration of trypsin or chymotrypsin inhibits cholecystokinin release and pancreatic secretion. The increased plasma cholecystokinin levels following diversion of pancreatic juice seems to be mediated by "cholecystokinin-releasing factor", a trypsin-sensitive substance secreted by the proximal small intestine. This factor may mediate pancreatic enzyme secretion in response to protein intake. Dietary protein in the intestine competes for the trypsin that would otherwise inactivate the factor. The resulting increase of this factor in the intestinal lumen releases cholecystokinin and stimulates pancreatic enzyme secretion. Enteropancreatic reflex can also be activated by distension or administration of hyperosmolar solutions in the duodenum eliciting pancreatic enzyme secretion without raising plasma cholecystokinin levels. This effect is inhibited by atropine, suggesting that it is cholinergically mediated. Pancreatic response to duodenal volume or osmolality is not suppressed by trypsin, indicating that the reflex is not affected by intraluminal proteases. Our studies also show that secretion of pancreatic polypeptide is under cholinergic control, and this peptide acts by interfering with cholinergic transmission, making it an ideal candidate to modulate pancreatic secretion stimulated by the vagal cholinergic pathway. Similar observations are made with somatostatin and calcitonin-gene related peptide which also acts preferentially to inhibit pancreatic secretion by the vagal cholinergic pathway.
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PMID:Negative feedback control of exocrine pancreatic secretion: role of cholecystokinin and cholinergic pathway. 791 21

Tryptase and chymase released from activated mast cells degrade the neuropeptides calcitonin gene-related peptide (CGRP) and vasoactive intestinal peptide (VIP) to peptide fragments. We have examined whether nedocromil sodium can modulate the ability of rat activated peritoneal mast cells to degrade 125I-CGRP and 125I-VIP. Mast cell-dependent degradation of both 125I-CGRP and 125I-VIP was observed with compound 48/80 (0.03-1 microgram/ml) and in the case of 125I-VIP with anti-IgE (1-20 micrograms/ml). Nedocromil sodium (10(-6)-10(-4) M) caused significant inhibition of neuropeptide degradation, with the most effective inhibition observed against anti-IgE-induced degradation of 125I-VIP. Nedocromil sodium had no inhibitory effect on the ability of lysed mast cells, bovine trypsin or chymotrypsin to breakdown 125I-VIP. These results suggest that nedocromil sodium inhibits mast cell-dependent degradation of neuropeptides, such as VIP, as a secondary consequence of inhibiting the release of mast cell proteases.
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PMID:The modulation by nedocromil sodium of proteases released from rat peritoneal mast cells capable of degrading vasoactive intestinal peptide and calcitonin gene-related peptide. 839 43

The purpose of this study was to determine whether carbopol polymers, polyacrylic acid polymers, can inhibit lumenal degradation of insulin, calcitonin and insulin-like growth factor I (IGF-I) by trypsin and chymotrypsin and to understand whether reducing the pH of the incubation medium by these polymers results in inhibition. Further, the effects of carbopol polymers on the in-situ absorption of insulin were studied in rats. In saline, carbopol polymers at 1% and 4% (w/v%) inhibited close to 100% of trypsin and chymotrypsin activities against insulin. In 50 mM Tris buffer, carbopol polymers, including 934P, 974P and 971P, at 0.1% only weakly inhibited degradation of calcitonin and insulin by both enzymes; however, as the polymer concentration increased to 0.4%, degradation of insulin, calcitonin, and IGF-I by both enzymes was complete or almost complete. When the Tris buffer was increased to 100 mM, no inhibition was observed at 0.1%. Determination of the final pH of the incubation medium in the presence of polymers revealed that the inhibitory effects of carbopol polymers correlated with the final pH. When the incubation medium has no or low buffer capacity to buffer the protons released by carbopol polymers, these polymers are able to reduce the pH much lower than the optimum pH for the enzyme activities, and thus inhibit proteolytic degradation. When the buffer capacity of the incubation medium increases, the inhibitory effects of carbopol polymers weaken. In-situ absorption of insulin revealed that carbopol polymers improved insulin absorption and induced a significantly greater decline in blood glucose levels. It is concluded that carbopol polymers with strong bioadhesive properties also can inhibit lumenal degradation of peptide hormones, offering multiple advantages for their uses in oral drug delivery.
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PMID:Effects of polyacrylic polymers on the degradation of insulin and peptide drugs by chymotrypsin and trypsin. 872 88

The objective of this study was to examine the metabolism of insulin and calcitonin, and their protection by various protease inhibitors, in the large intestine. Fresh caecal contents were prepared from non-fasted rats and the degradation of insulin and calcitonin was studied in a suspension of rat caecal contents, as a model of the content of the large intestine. Both insulin and calcitonin were metabolized in suspensions of rat caecal contents, but the degradation of calcitonin was much faster than that of insulin. The degradation of insulin was fastest at pH 6.8. Protease inhibitors such as camostat and aprotinin inhibited the degradation of insulin and calcitonin in rat caecal contents, which was consistent with the high chymotrypsin activity of these contents. These findings suggest that care should be taken when administering peptide drugs to the large intestine for colon-specific drug delivery because they can be degraded in rat caecal contents. Protease inhibitors might be useful for increasing the stability of these peptides in the large intestine, thereby improving their large-intestinal absorption to the systemic circulation.
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PMID:Degradation of insulin and calcitonin and their protection by various protease inhibitors in rat caecal contents: implications in peptide delivery to the colon. 905 89

The semisynthesis of eel[L-alpha-aminosuberic acid]calcitonin (elcatonin) was accomplished by alpha-chymotrypsin-catalyzed coupling of two peptide segments in a single reaction without the protection of any functional group. The eel calcitonin-(10-32)-peptide was prepared by a gene manipulation. The esters of cyclic desamino nonapeptide (segment 1-9) were synthesized by the conventional solution method including a thermolysin-mediated resolution of DL-alpha-aminosuberic acid via one-step tripeptide synthesis leading to the 7-9 sequence. The main aim of this work was to determine the conditions for protease-catalyzed segment condensation while avoiding a concurrent cleavage of other proteolytically labile peptide bonds in the hormone. The alpha-chymotrypsin condensation strategy under usual conditions led to a complicated mixture of split products with an insignificant amount of the required peptide. When the coupling reaction was carried out at 0 degrees C, the reaction resulted in a satisfactory yield of elcatonin with the complete conversion of the acyl donor (1-9 segment) accompanied by negligible concurrent peptide bond digestion. The same strategy was employed for the preparation of analogous dicarba salmon calcitonin using a synthetic elcatonin-(10-32)-peptide. Both calcitonin analogs exhibited hypocalcemic activity corresponding to the international standard of elcatonin. We demonstrate in this work a peptide synthesis based on the combination of genetic engineering, chemical synthesis and proteinase-catalyzed segment condensation. This approach enables effective incorporation of an unnatural amino acid into calcitonins without the side-chain protection.
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PMID:Enzymatic semisynthesis of dicarba analogs of calcitonin. 924 31

1. To investigate further the release, localization and identity of a non-nitrergic mediator of smooth muscle relaxation in the female pig urethra, we studied the effects of drugs acting at alpha 2-adrenoceptors or K+ channels, the effects of capsaicin and chemical sympathectomy, and the actions of several transmitter candidates. 2. Electrical field stimulation (EFS; frequencies above 12 Hz) of spontaneously contracted smooth muscle strips from the female pig urethra evoked long-lasting, frequency-dependent relaxations in the presence of prazosin, scopolamine, and NG-nitro-L-arginine. Treatment with the selective alpha 2-adrenoceptor agonist UK-14 304 markedly reduced the relaxations evoked by EFS at all frequencies tested (16-30 Hz). The inhibitory effect of UK-14 304 was completely antagonized by the alpha 2-adrenoceptor antagonist rauwolscine. The muscarinic M1 receptor antagonist, pirenzepine, or exogenously administered carbachol, did not have any effects on the electrically evoked relaxations. 3. Inhibition of high conductance Ca2+ activated K+ channels by iberiotoxin or charybdotoxin significantly enhanced the relaxations evoked by EFS at all frequencies. However, inhibition of voltage-sensitive K+ channels with 4-aminopyridine or dendrotoxin-1, treatment with the ATP-sensitive K+ channel blocker, glibenclamide, or treatment with the high and low conductance Ca2+ activated K+ channel blockers, tetraethylammonium chloride and apamin, had no effect on the relaxations evoked by EFS. 4. Electrically evoked relaxations were not affected by adrenergic denervation with 6-hydroxydopamine (6-OHDA) at any frequency. However, treatment with 6-OHDA abolished prazosin-sensitive electrically induced contractions, and a long-lasting relaxation was revealed. Treatment with capsaicin, believed to damage selectively a subpopulation of primary afferent fibres, did not affect basal tone or relaxations evoked by EFS. 5. Exogenously applied vasoactive intestinal polypeptide (VIP), pituitary adenylate cyclase-activating peptide (PACAP)-27, PACAP-38, adenosine, ATP and 5-hydroxy-tryptamine caused relaxations of the urethral preparations, whereas prostaglandin E2 and calcitonin gene-related peptide had no effects. VIP 10-28, alpha, beta-methylene-ATP, reactive blue-2, suramin or indomethacin did not reduce the electrically-evoked relaxations at any frequency. However, the relaxations were slightly reduced by trypsin or alpha-chymotrypsin. 6. The present results suggest that the release of the unknown mediator in the female pig urethra can be modulated via alpha 2-adrenoceptors and high conductance Ca2+ activated K+ channels. Furthermore, the mediator does not appear to be localized to or released from adrenergic or capsaicin-sensitive sensory nerve-endings. The identity of the transmitter remains to be established.
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PMID:NANC transmitters in the female pig urethra--localization and modulation of release via alpha 2-adrenoceptors and potassium channels. 928 93


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