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)

The amino acid sequence of the alpha-chain of the major haemoglobin of a newt, T. granulosa, has been determined. The chain is 142 residues long and has an extra methionine at its N-terminus when compared with human alpha-chain. Most of the tryptic peptides were sequenced by a combination of the subtractive Edman method and by deduction from the compositions of overlapping fragments produced by various enzymic treatments. The sequence of two 'core' regions was obtained by automatic sequencing of large peptides produced by trypsin cleavage at arginine residues only after blockage of lysine residues by citraconylation; by cleavage between aspartic acid and proline residues with 70% formic acid, and by cyanogen bromide cleavage at methionine residues. The sequence of T. granulosa alpha-chain is compared with those of representative species from the other classes of vertebrates. The differences in alpha-chain between the classes of vertebrates are compared with the differences in this protein between an equal number of orders of mammals. This comparison allows us to conclude that the major functional and conformational features of alpha-chain have been conserved since the divergence of the classes of jawed vertebrates.
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PMID:alpha-chain sequence of newt haemoglobin (Taricha granulosa). 90

Two isoinhibitors (II and III-B) have been isolated from kidney bean (Phaseolus vulgaris L.) in a highly purified state. Both were active against trypsin and chymotrypsin to the same extent. Their amino acid composition is characterized by a high content of half-cystine, aspartic acid (or asparagine) and serine, by the absence of valine, methionine and tryptophan. Glycine and serine were N-terminal in II and III-B respectively. Both isoinhibitors have C-terminal leucine.
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PMID:[Isolation and properties of trypsin isoinhibitors from kidney beans]. 90 94

The beta-chain of fibrin was cleaved with trypsin or cyanogen bromide. Carbohydrate containing fragments were isolated by affinity chromatography on concanavalin A-agarose. The fragments were analysed for amino acid sequence and for amino and carbohydrate composition. The sequence of 21 amino acid residues around the carbohydrate attachment point was determined. The carbohydrate carrying amino acid was identified as aspartic acid/asparagine.
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PMID:Carbohydrate linkage site in the beta-chain of human fibrin. 100 28

The monoclonal antibody, 3/4/2, which was raised against purified rat cytochrome P450 isoenzyme 1A1 (CYP1A1) binds to cytochromes P4501A in many species. It was shown by immunoblotting that the antibody binds to CYP1A1 in microsomal fractions prepared from rat, mouse, rabbit, hamster and human. The antibody also binds to cytochrome P450 isoenzyme 1A2 in microsomal fractions prepared from rabbit and human, but not rat or mouse. Using purified isoenzymes in an enzyme-linked immunosorbent assay it was found that the affinity of binding to the two rabbit hydrocarbon-inducible isoenzymes is reduced compared with that for rat CYP1A1. Binding is not affected by denaturation of the antigens. The effects of chemical and enzymatic treatments on rat CYP1A1 showed that the epitope contains a trypsin-sensitive site that includes arginine, but lacks lysine. The epitope does not contain methionine, cysteine, aspartic acid or glutamic acid residues. In addition, digestion of the protein with cyanogen bromide produces a fragment of Mr 20,000 which contains the antibody binding site. By comparing the cross-reactivity of the antibody with the primary structures of CYP1A1 and 1A2 from the rat, mouse, rabbit and human, and by considering the results of the chemical and enzymatic treatments, it was possible to deduce the likely location and structure of the binding site of 3/4/2 on members of the CYP1A subfamily. It is concluded that the epitope for this antibody is Phe-Arg-His-Ser-Ser-Phe, which lies at positions 380-385 in rat CYP1A1. Further, it is predicted from a model of the tertiary structure of eukaryotic cytochrome P450 that a part of this binding site lies within a helix in the native protein.
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PMID:Identification of the epitope of a monoclonal antibody which binds to several cytochromes P450 in the CYP1A subfamily. 137 49

We recently reported the purification of a lymphocyte granule protein called "fragmentin," which was identified as a serine protease with the ability to induce oligonucleosomal DNA fragmentation and apoptosis (Shi, L., R. P. Kraut, R. Aebersold, and A. H. Greenberg. 1992. J. Exp. Med. 175:553). We have now purified two additional proteases with fragmentin activity from lymphocyte granules. The three proteases are of two types; one has the unusual ability to cleave a tripeptide thiobenzyl ester substrate after aspartic acid, similar to murine cytotoxic cell protease I/granzyme B, while two are tryptase-like, preferentially hydrolyzing after arginine, and bear some homology to human T cell granule tryptases, granzyme 3, and Hanukah factor/granzyme A. Using tripeptide chloromethyl ketones, the pattern of inhibition of DNA fragmentation corresponded to the inhibition of peptide hydrolysis. The Asp-ase fragmentin was blocked by aspartic acid-containing tripeptide chloromethyl ketones, while the tryptase fragmentins were inhibited by arginine-containing chloromethyl ketones. The two tryptase fragmentins were slow acting and were partly suppressed by blocking proteins synthesis with cycloheximide in the YAC-1 target cell. In contrast, the Asp-ase fragmentin was fast acting and produced DNA damage in the absence of protein synthesis. Using a panel of unrelated target cells of lymphoma, thymoma, and melanoma origin, distinct patterns of sensitivity to the three fragmentins were observed. Thus, these three granule proteases make up a family of fragmentins that activate DNA fragmentation and apoptosis by acting on unique substrates in different target cells.
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PMID:Purification of three cytotoxic lymphocyte granule serine proteases that induce apoptosis through distinct substrate and target cell interactions. 146 Apr 16

Oversulphated chondroitin sulphate proteoglycan from squid skin was isolated from 4 M guanidine hydrochloride extract by ion-exchange chromatography, gel chromatography and density gradient centrifugation. The proteoglycan had Mr 3.5 x 10(5), contained on average six oversulphated chondroitin sulphate chains (Mr 4 x 10(4)) bound on a polypeptide of Mr 2.8 x 10(4), and oligosaccharides consisting of both hexosamines, glucuronic acid, sulphates and fucose as the only neutral monosaccharide. The major amino acids of the proteoglycan protein core are glycine (corresponding to about one third of the total amino acids), aspartic acid/asparagine and serine, together amounting to 50% of the total. The proteoglycan was resistant to the proteolytic enzymes V8 protease, trypsin (treated with diphenylcarbamoyl chloride), alpha-chymotrypsin and pronase, while it was completely degraded by papain and to a large extent by collagenase. Pretreated proteoglycan with chondroitinase AC was degraded by pronase to a large extent and slightly by V8 protease and trypsin. The proteoglycan did not interact with hyaluronic acid and did not form self-aggregates. Oversulphated chondroitin sulphate chains were composed of unusual sulphated disaccharide units which were isolated and characterized by HPLC. In particular, it contained 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid)-D-galactose 4-sulphate (delta di-4S) and disulphated disaccharides (delta di-diS) [90% 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid 2/3-sulphate)-D-galactose 6-sulphate (delta di-diSD) and 10% 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid 2/3-sulphate)-D-galactose 4-sulphate (delta di-diSK)] as the major disaccharides, significant amounts of trisulphated disaccharides (delta di-triS) and small amounts of 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid)-D-galactose 6-sulphate (delta di-6S) and 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid)-D-galactose (delta di-OS). Trisulphated disaccharides contained sulphate groups at C-4 and C-6 of the galactosamine and at C-2 or C-3 of the glucuronic acid. By HPLC analysis of a pure preparation of oversulphated chondroitin sulphate, it was found that it contains glucose, galactose, mannose and fucose most likely as branches.
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PMID:Isolation, characterization and properties of the oversulphated chondroitin sulphate proteoglycan from squid skin with peculiar glycosaminoglycan sulphation pattern. 154 Dec 70

Cytolytic granules purified from natural killer lymphocytes (NK) contain a pore-forming protein (perforin) and a number of serine proteases. When these proteases are inhibited by serine protease-specific isocoumarin reagents the serine proteases are inactivated and the cytolytic activity of the granules is decreased. Paradoxically, it has been found that the general serine protease inhibitor phenylmethylsulfonyl fluoride (PMSF) frequently cannot block killing even though it inhibits many of the serine proteases. At the same time it has been reported that "purified" perforin alone can lyze cells. To address these inconsistencies we first compared the ability of PMSF and four new sulfonyl fluoride serine protease inhibitors to inhibit proteases and cell lysis. We determined the effects on lysis and the second order inhibition rate constants for five granule protease activities: ly-tryptase, ly-chymase, Met-ase (methionine cleaving), Ser-ase (serine cleaving) and Asp-ase (aspartic acid cleaving). One compound, 2-(Z-NH(CH2)2CONH)C6SO2F, was a potent inhibitor of Met-ase activity (k(obsd)/[I] = 162 M-1 s-1), ly-chymase activity (k(obsd)/[I] = 147 M-1 s-1), and granule-mediated as well as perforin-mediated lysis. PMSF was a poor inhibitor of granule proteases (k(obsd)/[I]'s less than 7 M-1 s-1 for four activities and no inhibition of Ser-ase); the lack of reactivity is consistent with the failure of PMSF to block granule lytic activity. We also prepared enriched perforin by anion exchange chromatography and showed that a ly-chymase and a Met-ase associated with perforin. By inhibiting these proteases we also inhibited lytic activity.
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PMID:Sulfonyl fluoride serine protease inhibitors inactivate RNK-16 lymphocyte granule proteases and reduce lysis by granule extracts and perforin. 160 92

Following incubation of UMR-106 cells for 48 h in the presence of [3H]glucosamine and [35S]sulfate, the newly synthesized anionic glycoconjugates were isolated from the culture medium by cetylpyridinium chloride/ethanol precipitation and further separated by DEAE-Sephacel chromatography into two radiolabelled fractions, a major component, UM I, and a minor component, UM II. UM I appeared to be homogeneous as shown by Sepharose CL-4B chromatography under dissociative conditions, and SDS-polyacrylamide gel electrophoresis. It showed a molecular mass of approximately 93 kDa on 4-15% gels. UM I was partially degraded by brief treatment with trypsin, releasing a small, terminal peptide that contained 47.6% of 35S but no 3H. Treatment of UM I with neuraminidase and 0.1 N H2SO4 (1 h at 80 degrees C), respectively, released 27% 3H and 38.4% 3H plus 41% 35S, suggesting the presence of a significant number of sialic acid residues, as shown by Sephadex G-50 chromatography of the digests. Amino acid analysis showed that the UM I glycoconjugate was rich in acidic amino acids (12.6% aspartic acid and 21.2% glutamic acid residues) and its N-terminal sequence was Phe-Ser-Met-Lys-Asn-Phe-, which is identical to the published N-terminal amino acid sequence of rat bone sialoprotein II. Keratanase treatment of UM I released 26% of the incorporated radioactivity, suggesting the presence of keratan sulfate chains. UM II contained a chondroitinase ABC-sensitive proteoglycan.
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PMID:Further purification and characterization of newly synthesized anionic glycoconjugates secreted by cultured UMR-106 cells: evidence that the major anionic glycoconjugate secreted by these cells is similar to bone sialoprotein II. 176 Jan 56

A protease inhibitor has been purified by ultracentrifugation, affinity chromatography on trypsin-sepharose 4B, and chromatofocusing on PBE-94 from hemolymph of the scorpion Heterometrus bengalensis. Homogeneity of the protease inhibitor was demonstrated by high performance liquid chromatography (HPLC). The protease inhibitor is a monomeric glycoprotein with a molecular weight of 120,000 dalton, which is stable between pH 4 and pH 8. The molecule inhibits serine proteases like trypsin and alpha-chymotrypsin and shows a noncompetitive mode of inhibition towards trypsin, with a Ki value of 6.1 x 10(-6) mM. Amino acid analysis shows a preponderance of aspartic acid, glutamic acid, serine, and glycine. The protease inhibitor is efficient in inhibiting phenoloxidase activity in both the hemolymph and the isolated phenoloxidase. Melanin synthesis by phenoloxidase may be influenced by this protease inhibitor.
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PMID:Characterization of a naturally occurring protease inhibitor in the hemolymph of the scorpion, Heterometrus bengalensis. 177 46

A trypsin inhibitor (ACTI) was isolated and purified from the seeds of Acacia confusa by gel filtration, and trypsin-Sepharose 4B column affinity chromatography. The molecular weight of ACTI was found to be 21,000 +/- 1,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and amino acid composition analysis. ACTI contained four half-cystine and no methionine residues, and was rich in aspartic acid, glutamic acid, glycine, leucine, and lysine residues. The native trypsin inhibitor was composed of two polypeptide chains, and it inhibited trypsin and alpha-chymotrypsin stoichiometrically at the molar ratio of 1:1 and 2:1, respectively. The amino-terminal sequence analysis of the A. confusa trypsin inhibitor A and B chains revealed a more extensive homology with Acacia elata and silk tree trypsin inhibitors, and a less extensive homology with Kunitz soybean trypsin inhibitor.
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PMID:Trypsin inhibitor from the seeds of Acacia confusa. 179 77


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