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)

Despite advances in human islet isolations, there remain inconsistencies in human islet yield and viability after collagenase digestion. It has been suggested that trypsin may contribute to the proteolysis of collagenase and the destruction of islet cells, or possibly exert indirect effects on the pancreas by activating other endogenous serine proenzymes. This study evaluated the effects of serine proteases on collagenase activity and profiled the kinetics of serine protease activity throughout human islet isolations with and without addition of Pefabloc, a serine protease inhibitor. Cadaveric pancreases were perfused in the presence (n = 12) and absence of Pefabloc (0.4 mmole; n = 8). Samples were collected before and throughout the digestion process and were assayed for trypsin, chymotrypsin, and elastase activity. A study of the enzyme kinetics of serine proteases throughout human islet isolations showed an increase in activity levels throughout the digestion period. There was a significant difference in the chymotrypsin (1342 +/- 503 and 384 +/- 71 units) and elastase (7.94 +/- 1.1 and 2.761 +/- 0.69 units) levels between the control and Pefabloc-supplemented isolations, respectively. There was no significance difference noted among the trypsin (88 +/- 27 and 54 +/- 18 units) levels between the control and Pefabloc-supplemented isolations, respectively. This demonstrates that serine proteases are effectively inhibited by Pefabloc during the islet isolation process. These data show that the presence of serine proteases may likely damage the islets upon prolonged digestion of the pancreatic tissue.
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PMID:An evaluation of the activation of endogenous pancreatic enzymes during human islet isolations. 1461 84

Ovoinhibitor is a serine protease-inhibiting protein that was originally purified from egg whites. It is secreted by the oviduct under the control of estrogen and progesterone and it specifically inhibits serine proteinases such as trypsin and chymotrypsin. During recent attempts to raise monoclonal antibodies (Mabs) against chicken bursa of Fabricius proteins, one Mab was produced that specifically recognized chicken ovoinhibitor. This was the first demonstration of ovoinhibitor in an avian immune organ. We presently report on the expression of an ovoinhibitor-like molecule by the pituitary of the chicken as revealed by immunocytochemistry and RT-PCR. Immunofluorescent dual staining experiments using the mouse anti-ovoinhibitor Mab in conjunction with polyclonal antibodies against various hypophysial hormones revealed partial co-localization of an ovoinhibitor-like molecule with growth hormone (GH), luteinizing hormone (LH), and pro-opiomelanocortin (POMC), in a subset of the respective hormone producing cells. By contrast, no co-localization with prolactin (PRL) could be reliably demonstrated. RT-PCR of hypophysial mRNA using ovoinhibitor gene-specific primers yielded an amplicon that was 20% shorter than predicted on the basis of the published ovoinhibitor sequence. Sequencing revealed that of the represented exons only the central portion was expressed in the pituitary and that both 5' and 3' ends of each exon had been truncated. While expression of ovalbumin-like serine protease inhibitors (serpins) has been previously reported in the rat pituitary, to our knowledge, this is the first report of a Kazal-type serine protease inhibitor in the vertebrate neuroendocrine system.
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PMID:The chicken pituitary expresses an ovoinhibitor-like protein in subpopulations of some, but not all, hormone-producing cell types. 1465 38

A colorimetric method for serine protease inhibition was modified using N-Acetyl-DL-Phenylalanine beta-Naphthylester (APNE) as the substrate and o-Dianisidine tetrazotized (oD) as the dye. The reaction generated a single peak absorbing at 530 nm for both trypsin and chymotrypsin. Standard curves with increasing enzyme concentrations showed strong linearity. A standard curve for the serine protease inhibitor, Bowman-Birk Inhibitor (BBI), has been made using this modified method. The IC50 for 3 U of trypsin was found to be 33 ng and the IC50 obtained for 3 mU of chymotrypsin was 53 ng. A recombinant BBI (rBBI) gene was constructed, cloned and expressed in the yeast Pichia pastoris. Evaluating samples of rBBI for protease inhibitory activity by the gel activity method failed to quantify the inhibitor amounts, due to high sensitivity for trypsin inhibition and low sensitivity for chymotrypsin inhibition. After development, the results could not be quantified, even to the extent that 1 microl of rBBI could not be detected with chymotrypsin inhibition. Therefore, a modified method for trypsin and chymotrypsin inhibition was used to evaluate the level of rBBI-expression for these same samples. The level of rBBI expression was calculated to be 50-56 ng/microl of media. These amounts fit into the range of values previously obtained by Western blot analysis. This modified method allows us to combine the sensitivity of the gel activity method with the quantification attributes of a Western blot. Thus, the modified method represents a significant improvement in speed, sensitivity and reproducibility over the gel activity method.
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PMID:A simple method to determine trypsin and chymotrypsin inhibitory activity. 1516 55

Tumors have several mechanisms to escape from the immune system. One of these involves expression of intracellular anticytotoxic proteins that modulate the execution of cell death. Previously, we have shown that the serine protease inhibitor (serpin) SPI-6, which inactivates the cytotoxic protease granzyme B (GrB), is capable of preventing cytotoxic T lymphocyte (CTL)-mediated apoptosis. Despite its potent antiapoptotic activity, SPI-6 does not prevent membranolysis induced by cytotoxic lymphocytes. We now provide evidence that several colon carcinoma cell lines do resist membranolysis and that this protection is dependent on SPI-6 but also requires expression of a closely related serpin called SPI-CI (serine protease inhibitor involved in cytotoxicity inhibition). Expression of SPI-CI is absent from normal colon but observed in placenta, testis, early during embryogenesis, and in cytotoxic lymphocytes. SPI-CI encodes a chymotrypsin-specific inhibitor and irreversibly interacts with purified granzyme M. Moreover, SPI-CI can protect cells from purified perforin/GrM-induced lysis. Our data therefore indicate that SPI-CI is a novel immune escape molecule that acts in concert with SPI-6 to prevent cytotoxic lymphocyte-mediated killing of tumor cells.
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PMID:SPI-CI and SPI-6 cooperate in the protection from effector cell-mediated cytotoxicity. 1545 90

Alpha-1-antitrypsin (alpha1-AT) is a member of the serine protease inhibitor family regulating numerous proteolytic processes. The genetic disorder, alpha1-AT deficiency, is well known as a cause of hereditary pulmonary emphysema and liver cirrhosis. To create an animal model of human alpha1-AT deficiency, we disrupted the major murine isoform PI2, which is similar to human alpha1-AT and is one of 7 alpha1-AT isoforms found in the mouse. The ability of the serum to inhibit the activities of human leukocyte elastase (HLE) and human chymotrypsin (CYT) was significantly lower in heterozygous mice (alpha1-AT/PI2 -/+) than wild-type (alpha1-AT/PI2 +/+) mice (73.2% vs. 100% for HLE and 67.8% vs.100% for CYT, respectively; P<0.05). The distribution of genotypes among F(2) progeny was not in accordance with Mendelian distribution (P<0.01), as the percentages of wild-type, heterozygotes and homozygotes were 47.8%, 37.3% and 14.9%, respectively. Thus, it is likely that impairment of the protease inhibitor had a critical effect on fetus development. The alpha1-AT/PI2 deficient mouse will be a useful animal model for elucidating the function of alpha1-AT in fetal development, studying the mechanisms of chronic inflammatory disease and evaluating therapeutic candidates for the treatment of inflammatory disease.
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PMID:Disruption of the murine alpha1-antitrypsin/PI2 gene. 1551 92

A 5.6 kDa trypsin-chymotrypsin protease inhibitor was isolated from the tubers of the potato (Solanum tuberosum L cv. Gogu) by extraction of the water-soluble fraction, dialysis, ultrafiltration, and C18 reversed-phase high performance liquid chromatography. This inhibitor, which we named potamin-1 (PT-1), was thermostable and possessed antimicrobial activity but lacked hemolytic activity. PT-1 strongly inhibited pathogenic microbial strains, including Candida albicans, Rhizoctonia solani, and Clavibacter michiganense subsp. michiganinse. Automated Edman degradation showed that the N-terminal sequence of PT-1 was NH2-DICTCCAGTKGCNTTSANGAFICEGQSDPKKPKACPLNCDPHIAYA-. The sequence had 62% homology with a serine protease inhibitor belonging to the Kunitz family, and the peptide inhibited chymotrypsin, trypsin, and papain. This protease inhibitor, PT-1, was composed of polypeptide chains joined by disulfide bridge(s). Reduced PT-1 almost completely lost its activity against fungi and proteases indicating that disulfide bridge is essential for its protease inhibitory and antifungal activity. These results suggest that PT-1 is an excellent candidate as a lead compound for the development of novel oral or other anti-infective agents.
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PMID:Antimicrobial activity studies on a trypsin-chymotrypsin protease inhibitor obtained from potato. 1580 84

Skeletal muscle atrophy in response to a number of muscle wasting conditions, including disuse, involves the induction of increased protein breakdown, decreased protein synthesis, and likely a variable component of apoptosis. The increased activation of specific proteases in the atrophy process presents a number of potential therapeutic targets to reduce muscle atrophy via protease inhibition. In this study, mice were provided with food supplemented with the Bowman-Birk inhibitor (BBI), a serine protease inhibitor known to reduce the proteolytic activity of a number of proteases, such as chymotrypsin, trypsin, elastase, cathepsin G, and chymase. Mice fed the BBI diet were suspended for 3-14 days, and the muscle mass and function were then compared with those of the suspended mice on a normal diet. The results indicate that dietary supplementation with BBI significantly attenuates the normal loss of muscle mass and strength following unloading. Furthermore, the data reveal the existence of yet uncharacterized serine proteases that are important contributors to the evolution of disuse atrophy, since BBI inhibited serine protease activity that was elevated following hindlimb unloading and also slowed the loss of muscle fiber size. These results demonstrate that targeted reduction of protein degradation can limit the severity of muscle mass loss following hindlimb unloading. Thus BBI is a candidate therapeutic agent to minimize skeletal muscle atrophy and loss of strength associated with disuse, cachexia, sepsis, weightlessness, or the combination of age and inactivity.
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PMID:Attenuation of skeletal muscle atrophy via protease inhibition. 1597 55

An antifungal protein, AFP-J, was purified from tubers of the potato (Solanum tuberosum cv. L Jopung) by various chromatographic columns. AFP-J strongly inhibited yeast fungal strains, including Candida albicans, Trichosporon beigelii, and Saccharomyces cerevisiae, whereas it exhibited no activity against crop fungal pathogens. Automated Edman degradation determined the partial N-terminal sequence of AFP-J to be NH2-Leu-Pro-Ser-Asp-Ala-Thr-Leu-Val-Leu-Asp-Gln-Thr-Gly-Lys-G lu-Leu-Asp-Ala-Arg-Leu-. The partially sequence had 83% homology with a serine protease inhibitor belonging to the Kunitz family, and the protein inhibited chymotrypsin, pepsin, and trypsin. Mass spectrometry showed that its molecular mass was 13 500.5 Da. This protease inhibitor suppressed over 50% the proteolytic activity at 400 microg/mL. These results suggest that AFP-J is an excellent candidate as a lead compound for the development of novel antiinfective agents.
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PMID:Kunitz-type serine protease inhibitor from potato (Solanum tuberosum L. cv. Jopung). 1607 39

A 6.5 kDa serine protease inhibitor was purified by anion-exchange chromatography from the crude extract of the Inga umbratica seeds, containing inhibitor isoforms ranging from 6.3 to 6.7 kDa and protease inhibitors of approximately 19 kDa. The purified protein was characterized as a potent inhibitor against trypsin and chymotrypsin and it was named I. umbratica trypsin and chymotrypsin inhibitor (IUTCI). MALDI-TOF spectra of the IUTCI, in the presence of DTT, showed six disulfide bonds content, suggesting that this inhibitor belongs to Bowman-Birk family. The circular dichroism spectroscopy indicates that IUTCI is predominantly formed by unordered and beta-sheet secondary structure. It was also characterized, by fluorescence spectroscopy, as a stable protein at range of pH from 5.0 to 7.0. Moreover, this inhibitor at concentration of 75 microM presented a remarkable inhibitory activity (60%) against digestive serine proteases from boll weevil Anthonomus grandis, an important economical cotton pest.
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PMID:Purification of a 6.5 kDa protease inhibitor from Amazon Inga umbratica seeds effective against serine proteases of the boll weevil Anthonomus grandis. 1610

Secretory leukocyte protease inhibitor (SLPI), a potent serine protease inhibitor, has been shown to suppress macrophage responses to bacterial lipopolysaccharide (LPS). SLPI contains two topologically superimposable domains. Its C-terminal domain binds and inhibits target proteases. It is not clear whether SLPI's anti-protease function plays a role in the LPS-inhibitory action of SLPI. Four single amino acid substitution mutants of SLPI, M73G, M73F, M73E and M73K, were generated. Wild type SLPI is a potent inhibitor of chymotrypsin and elastase. Mutants M73G and M73F selectively lost inhibitory function towards chymotrypsin and elastase, respectively, whereas mutants M73K and M73E inhibited neither elastase nor chymotrypsin. Macrophage cell lines were established from RAW264.7 cells to stably express each SLPI mutant. Expression of the SLPI protease inhibition mutants suppressed NO and TNF production in response to LPS in a similar fashion as wild type SLPI. Expression of truncated forms of SLPI, containing only its N-terminus or its C-terminus, was similarly sufficient to confer inhibition of LPS responses. Thus, the LPS-inhibitory action of SLPI is independent of its anti-protease function.
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PMID:Suppression of macrophage responses to bacterial lipopolysaccharide (LPS) by secretory leukocyte protease inhibitor (SLPI) is independent of its anti-protease function. 1611 12


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