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)

An acid-stable, low molecular mass proteinase inhibitor, bronchial mucus proteinase inhibitor (BMPI), has been isolated from sputum and partially characterised. A single band with a modal molecular mass of 18 700 was observed following SDS-polyacrylamide gel electrophoresis. BMPI inhibited human leukocyte elastase, cathepsin G, trypsin and chymotrypsin, but not porcine pancreatic elastase. Although BMPI had a molecular mass close to the similarly isolated inhibitor of Girard et al. (Girard, F., Tournier, J.M., Polu, J.M. & Sadoul, P. (1980), Bull. Eur. Physiopathol. Respir. 16 (Suppl.) 237-245), and although it showed immunological cross reactivity to the low molecular mass inhibitor of Kramps et al. (Kramps, J.A., Franken, C., Meyer, C.J.L.M. & Dijkman, J.H. (1981) J. Histochem. Cytochem. 29, 712-719), it was found to have an amino-acid profile different to any previously described inhibitor. BMPI was detectable in bronchoalveolar lavage fluid collected from healthy and diseased human lungs. The median molar ratio of BMPI/alpha 1-proteinase inhibitor (alpha 1 PI) observed in these lavage samples was 0.7, which is generally higher than those derived from the data of other authors. This suggests that BMPI is a different protein to those previously described, although its exact relationship to other low molecular mass proteinase inhibitors remains to be determined.
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PMID:Acid-stable low molecular mass proteinase inhibitors in human lung lavage. 308 87

Methods were developed for the preparation of biotinylated trypsin of high specific activity. This was used as a probe for the detection of serine proteinase inhibitory proteins which had been separated by sodium dodecyl sulfate-polyacrylamide gradient slab gel electrophoresis and electroblotted to nitrocellulose. The method was extremely sensitive and specific and could detect 0.15 ng of the serine proteinase inhibitor, trasylol (0.023 pmol active inhibitor). The method was also widely applicable and could be used to detect a range of serine proteinase inhibitors in human serum with molecular weights of 50 to approximately 180 kDa, soybean trypsin inhibitor variants, and secretory leukocyte proteinase inhibitor extracted from human intervertebral disc and articular cartilage. The identity of several of the serine proteinase inhibitors detected using biotinylated trypsin was verified by Western blotting using specific antibodies. Under the conditions used, electrotransfer of trasylol was quantitative; densitometric examination of blots indicated that there was a linear relationship between the amount of active trasylol electrophoresed (1-50 ng) and the intensity of the blot obtained with biotinylated trypsin as probe, indicating that serine proteinase inhibitory proteins may also be quantified by this technique using densitometric scanning.
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PMID:The preparation and use of biotinylated trypsin in western blotting for the detection of trypsin inhibitory proteins. 785 68

Binding of the human recombinant secretory leukocyte proteinase inhibitor (SLPI) [native and with the methionyl residues at positions 73, 82, 94 and 96 of domain 2 oxidized to the sulfoxide derivative (Met(O) SLPI)] to bovine alpha-chymotrypsin (alpha-chymotrypsin) [native and with the Met192 residue converted to the sulfoxide derivative (Met(O) alpha-chymotrypsin)] as well as to native bovine beta-trypsin (beta-trypsin), which does not contain methionyl residues, has been investigated between pH 4.0 and 8.0, and between 10.0 degrees C and 30.0 degrees C, from thermodynamic and/or kinetic viewpoints. By increasing the number of oxidized methionyl residues present at the proteinase:inhibitor contact interface (from 0 to 3), the adducts investigated are increasingly destabilized and the relaxation time of the complexes into conformers less stable is enhanced. On the other hand, the selective oxidation of methionyl residues of SLPI and alpha-chymotrypsin, by reaction with chloramine T, does not affect the proteinase inhibition recognition mechanism. Therefore, even though conformational changes may occur in the conversion of native SLPI and native alpha-chymotrypsin to their Met(O) derivatives, a localized steric hindrance can be considered as the main structural determinant accounting for the reported results.
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PMID:Selective oxidation of methionyl residues in the human recombinant secretory leukocyte proteinase inhibitor. Effect on the inhibitor binding properties. 798 66

Airway inflammation is often associated with the infiltration of activated neutrophils and subsequent protease release. Although aiding in the digestion and phagocytosis of foreign proteins and microorganisms, neutrophil proteases can indiscriminately damage healthy lung tissue. In the conducting airway, proteases, particularly neutrophil elastase, are counter-balanced by several antiproteases, including secretory leukocyte protease inhibitor (SLPI). SLPI can be produced locally by a number of cells including the airway epithelial cell. To examine the effects of neutrophil granule components on SLPI transcript levels, airway epithelial cells were treated (up to 96 h) with elastase, other proteases, or enzymes isolated from human sputum. We found that neutrophil elastase increased SLPI transcript levels in primary and transformed human airway epithelial cells in a time- and dose-dependent manner. Other neutrophil products, such as cathepsin G, myeloperoxidase, and lysozyme, had little or no effect on SLPI transcript levels. However, two nonneutrophil proteases, trypsin and pancreatic elastase, also increased SLPI transcript levels at higher doses than that required of neutrophil elastase. Two inflammatory cytokines, tumor necrosis factor-alpha and interleukin-8, produced little or no effect on SLPI transcript levels. This study demonstrates one way in which SLPI is regulated, via a protease that it inhibits, neutrophil elastase.
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PMID:Neutrophil elastase increases secretory leukocyte protease inhibitor transcript levels in airway epithelial cells. 810 97

Human secretory leukocyte protease inhibitor (SLPI), a serine protease inhibitor found concentrated in secretory fluids, has been postulated to participate in the body's natural defense against infection by the human immunodeficiency virus type-1 (HIV-1) by affecting trypsin-like enzymes on the surface of target cells. SLPI was evaluated for potential antiviral activity against laboratory, clinical and monocytotropic strains of HIV-1 in human T-cell lines, peripheral blood lymphocytes and monocyte/macrophage cultures. SLPI was tested in a single cycle of infection assay and under conditions in which SLPI was preincubated both with target cells and with virus and then maintained during the virus-to-cell adsorption phase and throughout the entire culture period. However, SLPI did not exert anti-HIV activity under any experimental conditions, and mechanistic studies showed SLPI to have no inhibitory activity on HIV-1 binding, reverse transcriptase or protease. Thus, SLPI exhibited no suggestive anti-HIV-1 activity.
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PMID:Human immunodeficiency virus type-1 (HIV-1) replication is unaffected by human secretory leukocyte protease inhibitor. 873 5

We found a novel trypsin-like enzyme (tryptase) in sputum from patients with chronic airway diseases, and named this enzyme human airway tryptase (HAT). To clarify its physiological significance in the airway, we compared biochemical properties of purified HAT with those of purified lung mast cell tryptase (MCT). Studies with model peptide substrates showed that both the HAT and MCT preferentially cleaved the COOH-terminal side of arginine residues of certain peptides, but substrate specificities to nine synthetic model substrates of HAT differed from those of MCT. Effects of protease inhibitors on the two enzymes were examined at a concentration of 10 microM. Both the HAT and MCT were strongly inhibited by the trypsin inhibitors leupeptin, antipain, and aprotinin. An alpha-1-protease inhibitor inhibited HAT by 50%, but it did not inhibit MCT. In contrast, a secretory leukocyte protease inhibitor strongly inhibited MCT, but not HAT. Mucoid sputum from patients with chronic bronchitis contained much more HAT than MCT. These differences in biochemical properties between HAT and MCT indicate that they play different physiological roles in the airways.
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PMID:[Comparison of biochemical properties of human airway tryptase isolated from mucoid sputum with those of lung mast cell tryptase]. 874 34

Our previous studies have shown that some human cancer cell lines produce pancreatic trypsinogen, plasminogen, and tissue-type kallikrein. To understand the regulatory mechanism of these proteinases, serine proteinase inhibitors secreted by human glioblastoma cell line T98G were analyzed by gelatin reverse zymography with trypsin. The serum-free conditioned medium of T98G cells showed more than ten trypsin inhibitor bands ranging from 16 to 150 kDa in the reverse zymography. Major trypsin inhibitors were purified by trypsin-affinity chromatography. Analysis of their N-terminal amino acid sequences demonstrated that the purified inhibitors were identical to the secreted forms of amyloid protein precursors (APPs), tissue factor pathway inhibitor (TFPI), placental protein 5 (PP5)/TFPI-2, and secretory leukocyte proteinase inhibitor (SLPI). In addition, a novel 25-kDa trypsin-binding protein, tentatively named p25TI, was identified. p25TI showed weak inhibitory activity against trypsin in reverse zymography as compared with the other inhibitors. The secretion of multiple forms of serine proteinase inhibitors by human cancer cells raises the possibility that they might be involved in the abnormal growth of cancer cells.
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PMID:Purification and identification of a novel and four known serine proteinase inhibitors secreted by human glioblastoma cells. 888 27

A novel trypsin-like protease was purified to homogeneity from the sputum of patients with chronic airway diseases, by sequential chromatographic procedures. The enzyme migrated on SDS-polyacrylamide gel electrophoresis to a position corresponding to a molecular weight of 28 kDa under both reducing and non-reducing conditions, and showed an apparent molecular weight of 27 kDa by gel filtration, indicating that it exists as a monomer. It had an NH2-terminal sequence of Ile-Leu-Gly-Gly-Thr-Glu-Ala-Glu-Glu-Gly-Ser-Trp-Pro-Trp-Gln-Val-Ser-Leu- Arg-Leu, which differed from that of any known protease. Studies with model peptide substrates showed that the enzyme preferentially cleaves the COOH-terminal side of arginine residues at the P1 position of certain peptides, cleaving Boc-Phe-Ser-Arg-4-methylcoumaryl-7-amide most efficiently and having an optimum pH of 8.6 with this substrate. The enzyme was strongly inhibited by diisopropyl fluorophosphate, leupeptin, antipain, aprotinin, and soybean trypsin inhibitor, but hardly inhibited by secretory leukocyte protease inhibitor at 10 microM. An immunohistochemical study indicated that the enzyme is located in the cells of the submucosal serous glands of the bronchi and trachea. These results suggest that the enzyme is secreted from submucosal serous glands onto the mucous membrane in patients with chronic airway diseases.
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PMID:Purification, characterization, and localization of a novel trypsin-like protease found in the human airway. 907 Jun 15

The major physiological role of human secretory leukocyte protease inhibitor (SLPI), a low molecular weight inhibitor present in mucus, is the rapid formation of a tight-binding inhibitory complex with neutrophil elastase. It is also the most effective known inhibitor of human mast cell chymase. The inhibitory efficacy of recombinant SLPI towards three other mast cell chymases was therefore investigated. Rat mast cell proteinases-1 and -2 (rMCP-1 and -2, respectively) and sheep mast cell proteinase-1 (sMCP-1), a chymase with additional tryptase-like properties, were treated with the inhibitor. SLPI inhibited rMCP-1 very efficiently in the absence of heparin, with a low dissociation constant, Ki = 3 x 10(-10) M and high second order association constant, kass = 8.0 x 10(6) M(-1) s(-1), and inhibition was enhanced when heparin was present. rMCP-2 was not inhibited by SLPI in the presence or absence of heparin, and did not degrade SLPI on prolonged incubation. SLPI inhibited sMCP-1 very poorly in the absence of heparin (Ki = 9 X 10(-6) M). However, in the presence of heparin, the Ki for inhibition of sMCP-1 by SLPI was reduced to the nanomolar range. sMCP-1 was observed to cleave SLPI with chymase-like specificity at Leu72-Met73 on prolonged incubation in the absence of heparin, but increasing concentrations of heparin reduced the extent of cleavage.
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PMID:Differential inhibition of mast cell chymases by secretory leukocyte protease inhibitor. 946 29

From purulent cystic fibrosis (CF) sputum, previous investigators partially purified a trypsinlike protease. A similar purified enzyme is available commercially as "human sputum trypsin." To explore the nature and origin of this preparation, we purified and NH2 terminally sequenced its major protein component. The resulting sequence, Ile-Val-Gly-Gly-Tyr-Thr-(Cys)-Ala-Ala-Asn-Ser-Val/Ile-Pro-Tyr-Gln-Val -Ser-Leu-Asn-Ser, differs from known human proteins but is identical to porcine trypsin, including the Val/Ile polymorphism at residue 12. Specific activity and electrophoretic and inhibition profiles and immunoreactivity of sputum and porcine pancreatic trypsin are nearly identical. Because porcine trypsin is a major ingredient of digestive enzyme supplements taken by CF patients with pancreatic dysfunction, we propose that one or more lots of human sputum trypsin derive from enzyme supplements and are of porcine origin. The path by which trypsin ends up in sputum is unknown. Because sputum trypsin is active but susceptible to inactivation by plasma alpha1-proteinase inhibitor, it is unlikely to derive from trypsin absorbed into the bloodstream. However, it may originate from tracheally aspirated stomach contents or from digestive supplement-contaminated saliva mixed with expectorated sputum. The imbalance between proteases and antiproteases in CF bronchial secretions allows trypsin to remain active despite sensitivity to serpins and secretory leukocyte proteinase inhibitor. Furthermore, because sputum trypsin activates human progelatinase B, it may be responsible in part for the reported presence of activated matrix metalloproteinases in CF sputum.
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PMID:Porcine origin of human sputum trypsin? 968 52


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