Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P15088 (mast cell)
 14,925 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The primary subsite specificities of human leukocyte elastase, cathepsin G, porcine pancreatic elastase, rat mast cell proteases I and II, bovine chymotrypsin A alpha, and the protease from strain V-8 of Staphylococcus aureus have been mapped with a series of tripeptide thiobenzyl ester substrates of the general formula Boc-Ala-Ala-AA-SBzl, where AA represents one of 13 amino acids. In addition, the effects of a P2 Pro and P4 methoxysuccinyl and succinyl groups were investigated. In an attempt to introduce specificity and/or reactivity into the substrate Boc-Ala-Ala-Leu-SBzl(X), the 4-chloro-, 4-nitro-, and 4-methoxythiobenzyl ester derivatives were studied. Enzymatic hydrolyses of the substrates were measured in the presence of 4,4'-dithiobis(pyridine) or 5,5'-dithiobis(2-nitrobenzoic acid), which provided a highly sensitive assay method for free thiol. The thio esters were excellent substrates for the enzymes tested, and in many cases, the best substrates reported here have kcat/KM values higher than those reported previously. The best substrate for human leukocyte elastase was Boc-Ala-Pro-Nva-SBzl(Cl), which has a kcat/KM of 130 X 10(6) M-1 s-1. A very reactive rat mast cell protease substrate, Boc-Ala-Ala-Leu-SBzl(NO2), was also found. The S. aureus V-8 protease was the most specific enzyme tested since it hydrolyzed only Boc-Ala-Ala-Glu-SBzl. Substituents on the thiobenzyl ester moiety of Boc-Ala-Ala-Leu-SBzl resulted in decreased KM values with human leukocyte elastase and rat mast cell protease I when compared to the unsubstituted derivative.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Active site mapping of the serine proteases human leukocyte elastase, cathepsin G, porcine pancreatic elastase, rat mast cell proteases I and II. Bovine chymotrypsin A alpha, and Staphylococcus aureus protease V-8 using tripeptide thiobenzyl ester substrates. 638 May 80

A flavin peptide from Corynebacterium sarcosine oxidase was obtained by proteolytic digestion with trypsin and chymotrypsin, and purified by the method of Kenney et al. (1). Amino acid analyses of the flavin peptide gave the following results: Asx(1), Ala(1), Val(1), and His(1) per flavin group. By carboxypeptidase A digestion and partial acid hydrolysis, the structure of the flavin peptide was determined as (Formula: see text).
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PMID:Amino acid sequence around the covalently-bound flavin prosthetic group of Corynebacterium sarcosine oxidase. 667 34

The formation of most connective tissue polysaccharides is initiated by transfer of D-xylose from UDP-D-xylose to specific serine residues in the core proteins of the putative proteoglycans. The substrate specificity of the xylosyltransferase catalyzing this reaction has not yet been examined in detail, but it appears that a -Ser-Gly- pair is an essential part of the substrate structure. Since the preparation of the known acceptors (e.g., Smith-degraded or HF-treated cartilage proteoglycan) involves a substantial effort, we have searched for readily available proteins with the -Ser-Gly-sequence, which might serve as alternative substrates. In the present work, it was found that silk fibroin from Bombyx mori, which consists, in large part, of the repeating hexapeptide, Ser-Gly-Ala-Gly-Ala-Gly, is an excellent substrate for the xylosyltransferase from embryonic chick cartilage. Pieces of silk were used directly in the reaction mixtures, and [14C]xylose transferred from UDP-D-[14C]xylose was measured by liquid scintillation spectrometry after rinsing the silk in 1 M NaCl and water. Substantially greater incorporation was observed with preparations of silk or fibroin which had been dissolved in 60% LiSCN and subsequently dialyzed exhaustively or diluted appropriately. Under standard reaction conditions, the Vmax for fibroin was 531 pmol/h/mg enzyme protein, as compared to 223 pmol/h/mg for Smith-degraded proteoglycan. Km values were 182 mg/liter (fibroin) and 143 mg/liter (Smith-degraded proteoglycan). The product of [14C]xylose transfer to silk was alkali labile, and [14C]xylitol was formed when [14C]xylosylsilk was treated with borohydride in alkali. Proteolytic digestion with papain, Pronase, leucine aminopeptidase, and carboxypeptidase A yielded a radioactive product which was identified as [14C]xylosylserine by electrophoresis and chromatography. The identity of the isolated [14C]xylosylserine was further supported by its resistance to treatment with alkali (0.5 M KOH; 100 degrees C; 8 h) and by acid hydrolysis which yielded [14C]xylose. Tryptic and chymotryptic fragments from fibroin were also good xylose acceptors and had Vmax values 60-70% of that observed for the intact protein. Substantial acceptor activity was displayed also by the sericin fraction of silk and by the silk sequence hexapeptide. Ser-Gly-Ala-Gly-Ala-Gly; the latter had a Vmax value close to 20% of that of intact fibroin.
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PMID:Silk--a new substrate for UDP-d-xylose:proteoglycan core protein beta-D-xylosyltransferase. 673 31

Multiple forms of urotensin II (UII), one of the hormonal peptides of the caudal neurosecretory system of fishes, were purified from the urophyses of the carp, Cyprinus carpio. Three distinct peaks with UII activity (classified as UII-alpha, -beta and -gamma) were separated by reverse-phase high-pressure liquid chromatography (HPLC). Edman degradation as well as digestion with carboxypeptidase A revealed the primary structures of these peptides as UII-alpha: Gly-Gly-Gly-Ala-Asp-Cys-Phe-Trp-Lys-Tyr-Cys-Val UII-beta: Gly-Gly-Ser-Asn-Thr-Glu-Cys-Phe-Trp-Lys-Tyr-Cys-Val UII-gamma: Gly-Gly-Gly-Ala-Asp-Cys-Phe-Trp-Lys-Tyr-Cys-Ile The results of thin-layer chromatography, HPLC, amino acid analysis, and sequencing indicate that UII-alpha and -gamma are homogeneous. UII-beta appears, however, to be a mixture of two components, differing only at position 2. Thus, in the carp urophysis, four forms of UII appear to be present, although the separation of two components in UII-beta has not been obtained. Sequence of positions 6-11 is common to all forms of UII isolated from the carp, sucker (Catostomus commersoni) and goby (Gillichthys mirabilis).
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PMID:Primary structures of multiple forms of urotensin II in the urophysis of the carp, Cyprinus carpio. 674 27

The phosphate transport protein from beef heart mitochondria has been purified on a large scale by hydroxylapatite chromatography in the presence of sodium dodecyl sulfate and urea. As shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (silver stain), the pure phosphate transport protein preparation consists of two protein bands (alpha and beta, ratio 1:1) with similar mobilities (34 kDa) which display identical peptide maps if fragmented with either CNBr or HCl/dimethyl sulfoxide/HBr. The complete amino acid composition of phosphate transport protein is presented. Quantitative determination of N-terminal amino acids underlines the purity of the preparation and shows for alpha and beta the identical amino-terminals H2N-Ala-Val-Glu-Glu-Glx-Tyr-. Qualitative digestion shows that carboxypeptidase A is able to release at least three amino acids from the C termini of the alpha as well as the beta band of phosphate transport protein. The nature of these two protein bands is discussed. The sum of phosphate transport protein (alpha + beta) per total mitochondrial protein amounts to 2.3% or 1.4 nmol of phosphate transport protein (34 kDa) per nmol of cytochrome b.
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PMID:Mitochondrial phosphate transport. Large scale isolation and characterization of the phosphate transport protein from beef heart mitochondria. 674 42

Carboxypeptidase A gamma from porcine pancreas was purified to homogeneity by ammonium sulfate fractionation, autolysis, batch absorption and elution from DEAF-Sephadex, and crystallization. The overall purification was about 32-fold with a yield of 31% and the specific activity of the purified protein was 108 units/mg protein. The apparent relative molecular mass determined by gel filtration on a Sephadex G-200 column was 38 900. The amino-terminal sequence of the porcine carboxypeptidase A gamma was Asn-Tyr-Ala-Thr-Tyr-His-Thr-Leu-Glu-Glu-Ile-Tyr-Asp-Phe-Met-Asp-Ile-Leu-Val-Ala -Glu-His-Pro-Gln-Leu- which was highly homologous to that of bovine carboxypeptidase A gamma. The purified enzyme was characterized with respect to isoelectric point (4.3). Km for N alpha-carbobenzoxyglycyl-L-phenylalanine (Cbz-Gly-LPhe) (20 mM), amino acid composition, pH optimum, pH stability, stability at different temperatures and effect of drying. The enzyme contained 1.01 mol zinc/mol and was inhibited by chelating agents such as EDTA and o-phenanthroline. Among substrates such as Cbz-Gly-LPhe, N alpha-benzoylglycyl-L-arginine, various kinds of amino acid esters, casein and elastin, porcine carboxypeptidase A gamma showed an enzymatic activity only towards Cbz-Gly-LPhe and casein. These data are in good agreement with the substrate specificity of bovine carboxypeptidase A.
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PMID:Crystallization and properties of carboxypeptidase A gamma from porcine pancreas. 727 15

Certain mast cell products, including the tetrapeptides Ala-Gly-Ser-Glu (Ala4) and Val-Gly-Ser-Glu (Val4) of the eosinophil chemotactic factor of anaphylaxis (ECF-A) and histamine, are preferentially chemotactic for eosinophils in vitro. Amino acid substitutions or modifications of the ECF-A tetrapeptides may substantially alter the chemotactic activity of human eosinophils. The cellular mechanism whereby ECF-A tetrapeptides enhance IgG-dependent killing of schistosomula by rat eosinophils is, however, unclear. We now report that the parent tetrapeptides and the substituted analogue Val-Pro-Ser-Glu (Pro3), but not histamine, increase the number of rat and human eosinophils that form rosettes with erythrocytes bearing IgG antibodies. Moreover, we find a correlation between the effect of these substances on the expression of rat eosinophil IgG Fc receptors and their capacity to increase the IgG-dependent cytotoxicity of rat eosinophils for Schistosoma mansoni schistosomula.
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PMID:Tetrapeptides of the eosinophil chemotactic factor of anaphylaxis (ECF-A) enhance eosinophil Fc receptor. 745 10

Cathepsin B activity was demonstrated histochemically in unfixed cryostat sections of inflamed human gingiva using the 2-methoxy-4-naphthylamide (MNA) substrates Z-Val-Lys-Lys-Arg-MNA and Z-Ala-Arg-Arg-MNA with a post-azo-coupling technique. Enzyme localisation was confirmed by immunocytochemistry with polyclonal sheep anti-human cathepsin B. In both cases, staining was found in connective tissue fibroblasts and also in cells varying in shape from rounded to more irregular forms. The latter were present both in areas of cellular infiltration and in the oral and pocket epithelium. Examination of adjacent sections with monoclonal antibodies directed against leukocyte differentiation antigens showed that the rounded to irregular cells were CD68 positive macrophages and monocytes. The histochemical staining had the form of fine cytoplasmic particles consistent with the known lysosomal occurrence of cathepsin B. Cells stained by the post-coupling method using the tryptase substrates Z-Ala-Ala-Lys-MNA and D-Val-Leu-Arg-MNA showed a different distribution and morphology, with reaction product confined to mast cell granules. The differences between the cathepsin B and tryptase staining patterns were confirmed by differential extraction from cryostat sections with salt-free and high-salt buffers respectively. Biochemical characterisation of activities in the extracts with the 7-amino-4-trifluoromethyl coumarin (AFC) substrates Z-Val-Lys-Lys-Arg-AFC and Z-Ala-Ala-Lys-AFC and protease inhibitors confirmed the identity of the two enzymes. Selective inhibitors could also be used in histochemical incubations to distinguish between cathepsin B and tryptase staining.
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PMID:Comparative histochemical, biochemical and immunocytochemical studies of cathepsin B in human gingiva. 751 62

Chymase, the major chymotryptic proteinase of human mast cells, can be released in substantial quantities following mast cell activation. As this enzyme is stored in the secretory granules in its fully active form, we have investigated various factors which might regulate its activity in storage and upon release. Chymase was purified from human skin by high salt extraction, cetylpyridinium chloride precipitation, heparin agarose affinity chromatography and gel filtration. Neither the addition of Mg2+ or Ca2+ (0.3-10 mM) nor their sequestration by EDTA had any effect on the rate of cleavage of the synthetic substrate N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide. Monovalent cations (Na+,K+) enhanced enzyme activity, but only at non-physiological concentrations (0.5-3.0 M), suggesting an ionic strength effect. At constant I = 0.15, enzyme activity was strongly pH-dependent: at pH 5.5 (the approximate pH of the mast cell granule) the activity was only 10% of that at pH 7.5 (the approximate pH of the extracellular space). Heparin, which is stored with chymase in the mast cell granule, accentuated this difference by enhancing activity at pH 7.5 by 33% and depressing it a pH 5.5 by 40%. Histamine at concentrations up to 50 mM (I = 0.15) had little effect on chymase activity at either pH, although high concentrations did attenuate the actions of heparin. It is concluded that pH and the interaction with heparin are central to the regulation of chymase activity within the granule and following release.
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PMID:Regulation of the activity of human chymase during storage and release from mast cells: the contributions of inorganic cations, pH, heparin and histamine. 761 63

Tryptase-like activity has previously been identified biochemically in gingival homogenates and gingival crevicular fluid (GCF) using substrates linked to the 7-amino-4-trifluoromethyl coumarin (AFC) leaving group. In the present study, activity was demonstrated histochemically in tissue sections with analogous 4-methoxy-2-naphthylamide (MNA) substrates. Z-Ala-Ala-Lys-MNA and D-Val-Leu-Arg-MNA were the most sensitive substrates. Comparison of staining patterns with the MNA substrates and toluidine blue indicated that enzyme activity was localized to mast cell secretory granules. Most stained cells were in the lamina propria, but a few were in the epithelium. The number of stained cells was somewhat greater in inflamed tissue from chronic periodontitis patients than in healthy tissue from controls. However, hardly any staining was seen in inflamed granulomatous tissue. Using high-salt buffer containing heparin, it was possible to extract enzyme activity from tissue sections for biochemical analysis with corresponding AFC substrates. Inhibitors gave similar results in the biochemistry and histochemistry. The inhibitor response and pH profile of the enzyme were the same as that found earlier with gingival homogenates and GCF and were again consistent with mast cell tryptase. The enzyme may have a role in the pathology of chronic periodontitis.
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PMID:Comparative histochemical and biochemical studies of mast cell tryptase in human gingiva. 769 9


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