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)

Mytilus pedal ganglia extract was treated sequentially with TPCK-trypsin and carboxypeptidase B. The treated sample and an untreated sample were purified separately with Sep-Pak C18 and subjected to HPLC. Fractions with Rt's corresponding to the Met-, Leu-enkephalin and Met-enkephalin-Arg6-Phe7 were assayed for enkephalin activities by displacement studies using pedal ganglia membrane and 3H-DAMA. The data showed the activities in the regions of Met-, Leu-enkephalin and Met-enkephalin-Arg6-Phe7 of the treated sample increased over that of the untreated sample by 2.3, 2.5 and 1.6 folds respectively. These results provided strong evidence for the presence of enkephalin precursor material in Mytilus.
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PMID:Presence of enkephalin precursor in molluscan neural tissue extract. 312 41

We have determined the partial amino acid sequences of the 40 kDa protein, one of the three pertussis toxin substrates in porcine brain. Purified 40 kDa protein from porcine brain was completely digested with TPCK-trypsin. Digested peptides were separated by reverse-phase HPLC and subjected to analysis by gas-phase protein sequencing. Several sequences of porcine brain 40 kDa protein completely matched with those which were deduced from the nucleotide sequences of the human Gi2 alpha gene and rat Gi2 alpha cDNA. On the other hand, the previously determined sequences of the rat brain 41 and 39 kDa proteins were in complete agreement with the predicted amino acid sequences of rat Gi1 alpha and Go alpha cDNAs, respectively.
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PMID:Identification of three pertussis toxin substrates (41, 40 and 39 kDa proteins) in mammalian brain. Comparison of predicted amino acid sequences from G-protein alpha-subunit genes and cDNAs with partial amino acid sequences from purified proteins. 312 41

Chymotrypsin can completely solubilize insoluble [3H]-labeled ligamentum nuchae elastin. At similar enzyme levels, trypsin solubilizes only 5% of the elastin substrate whereas pancreatic elastase completely solubilizes the elastin at one-tenth the concentration required for chymotrypsin solubilization. The elastolytic activity of chymotrypsin is dependent on Ca+2, is enhanced by SDS, and is inhibited by NaCl at concentrations above 10 mM. The elastolytic activity of chymotrypsin is also inhibited by TPCK, a chymotrypsin specific inhibitor, but not by TLCK, a trypsin specific inhibitor. Neither TPCK nor TLCK abolish the elastolytic activity of pancreatic elastase. The sizes of [3H]elastin fragments produced by the elastolytic activity of chymotrypsin are similar to those produced by pancreatic elastase, and larger than those produced by trypsin.
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PMID:Studies on the elastolytic activity of chymotrypsin. 316 94

Incubation at 37 degrees C of human cationic trypsinogen purified by PAGE electrophoresis, results in development of proteolytic activity (enzyme Y) capable of rapidly degrading cationic and anionic trypsinogens to inert products. Enzyme Y appears to be a serine protease with a molecular weight of about 20,000 daltons and is different from any of the known pancreatic enzymes. The active enzyme may be derived from trypsinogen itself or a hitherto unrecognized precursor contaminating the trypsinogen fraction used in this work. Appearance of enzyme Y activity seems to be associated with the presence of traces of free trypsin. Enzyme Y possesses insignificant or no activity when tested with a variety of synthetic trypsin, chymotrypsin and other protease substrates. It is not inactivated by the specific trypsin and chymotrypsin inhibitors TLCK and TPCK, but its activity is reduced gradually by increasing concentrations of pancreatic secretory trypsin inhibitor. Ca2+ concentrations greater than 3 mM strongly inhibit enzyme Y, and diisopropylfluorophosphate completely inactivates it. The enzyme is stable when incubated at pH 1.9 and 37 degrees C for 30 min and its activity is not abolished by treatment with Hg2+. When added to pancreatic juice with low inhibitor content it causes rapid inactivation of zymogens without significant release of active enzymes or reduction of pancreatic trypsin inhibitor. Its physiological role may be perceived as a second line of defense against premature intrapancreatic activation of zymogens. Enzyme Y activity may be generated when trypsin inhibitor, the first line of defense, is sufficiently depleted by complex formation with inappropriately released trypsin to permit dissociation of a small amount of trypsin from this complex. This in turn may lead to activation of enzyme Y and inactivation of the zymogens of pancreatic proteases.
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PMID:A possible zymogen self-destruct mechanism preventing pancreatic autodigestion. 316 6

We have used a polyclonal affinity-purified antibody made against chicken brain fodrin (both 240 and 235 Kd subunits) as a probe to determine if a fodrinlike protein exists in amoebae of Dictyostelium discoideum. In Western blots of whole cells and the isolated cell cortex, polypeptides measuring 220 and 70 Kd are recognized by the fodrin antibodies. In situ localization by indirect immunofluorescence with antifodrin indicates that the immunoreactive polypeptides are cortical. The immunoreactive analogues copatch and cocap with concanavalin A. At the level of resolution of the electron microscope, immunocytochemistry with antifodrin and colloidal gold confirms that the immunoreactive analogues are cortical proteins associated with microfilaments on the cytoplasmic side of the plasma membrane. We have isolated and characterized the 220 Kd protein to determine if it is similar to fodrin and to investigate its relationship to the 70 Kd polypeptide. The 220 Kd protein can be extracted from the cortex in the absence of detergent and isolated by gel filtration and sucrose density gradient sedimentation. The 220 Kd is a rod-shaped protein 118 +/- 17.8 nm (N = 37) in length. It has a sedimentation coefficient of 9.3 S and Stokes' radius of 13 nm and exists as a dimer of approximately 500,000 daltons (Mr). Isolated 220 Kd binds to actin filaments in vitro when assayed by rotary shadowing. Morphological criteria distinguish 220 Kd from Dictyostelium myosin II heavy chain (215 Kd) and the filaminlike protein at 240 Kd. The 70 Kd polypeptide appears to be a cleavage fragment of the 220 Kd, since it is found after prolonged storage when formerly only the 220 Kd was present. Furthermore, the 220 and 70 Kd polypeptides exhibit similar one-dimensional peptide maps when treated with TPCK trypsin. On the basis of its physical and immunoreactive characteristics, and location in the cell, the 220 Kd may be a fodrinlike protein.
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PMID:Isolation of an immunoreactive analogue of brain fodrin that is associated with the cell cortex of Dictyostelium amoebae. 321 34

A chymotrypsin-like enzyme (CTLE) was isolated from the digestive tract of the African migratory locust Locusta migratoria migratorioides by ion-exchange chromatography on diethylaminoethyl (DEAE) cellulose followed by affinity chromatography on phenylbutylamine (PBA) Sepharose. The purity and homogeneity of CTLE have been shown by SDS-PAGE and on cellulose acetate strips. The enzyme has a molecular weight of 24,000, determined by SDS-PAGE and on a Sephadex G-75 calibrated column. It has an isoelectric point of 10.1 and contains 0-1 half cystine residues. Sequence analysis of the first 20 N-terminal amino acids has shown 25% homology with bovine chymotrypsin and 40% homology with Vespa crabo and Vespa orientalis chymotrypsins and with Hypoderma lineatum trypsin. The optimal pH for enzyme activity and stability was in the range of 8.5-9.0. The Km and kcat values, determined on substrates for proteolytic, esterolytic and amidolytic activity, similar to those for bovine chymotrypsin. CTLE was inactivated by PMSF and TPCK indicating the involvement of serine and histidine in its active site. The enzyme was fully inhibited by the proteinaceous, double-headed, chymotrypsin-trypsin inhibitors BBI from soybeans and CI from chickpeas, by chicken ovomucoid (COM) and turkey ovomucoid (TOM), as well as by the Kunitz soybean trypsin inhibitor (STI) which hardly inhibits bovine chymotrypsin. Inhibition studies of CTLE with amino acid and peptide-chloromethylketones point towards the existence of an extended binding site.
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PMID:Purification and characterization of Locusta migratoria chymotrypsin. 324 83

Conditions for extraction of rat brain soluble and particulate cysteine proteinase inhibitors (CPIs) were compared and an optimal one was selected to isolate low- and high-molecular-weight forms active toward papain or brain cathepsins B/L. The different forms were purified by affinity chromatography on alkylated papain, and identified on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels by use of Schiff's reagent, or by immunoblots using antisera to monomer or polymeric forms of human urinary cystatin c, to a human plasma histidine-rich glycoprotein (HRG), or to rat plasma T-kininogen. In particulates containing nuclei (P1) or synaptosomes (P2) the predominant CPI was an 80-kDa glycoprotein cross-reacting to anti-HRG and shown to be a T-kininogen by treatment with TPCK-trypsin, and subsequent bioassay of the released kinins. The levels found in rat brain were approximately 0.5 nmol/g wet weight. The higher-molecular-weight CPI potently inhibited cathepsin L hydrolysis of Leu-enkephalin at the Gly2-Gly3 bond with a Ki 10(-10) M. In contrast the low-molecular-weight CPIs were present in postmicrosomal fractions (S3) and cross-reacted with anti-cystatin c, but not with anti-HRG, anti-lysozyme, anti-beta protein amyloid peptide, or anti-T-kininogen. The low-molecular-weight forms were present at approximately 1-1.5 nmol/g wet weight and resembled "cerebrocystatin" purified previously from rat brain cytosol by M. Kopitar, F. Stern, and N. Marks [1983) Biochem. Biophys. Res. Commun. 112, 1000-1006.).
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PMID:Diversity of rat brain cysteine proteinase inhibitors: isolation of low-molecular-weight cystatins and a higher-molecular weight T-kininogen-like glycoprotein. 326 47

Previously we have studied the binding domains on von Willebrand factor (vWF) involved in ristocetin-induced binding to platelets (ristocetin binding domain, RBD) and in the binding of vWF to collagen (collagen binding domain, CBD) using tryptic fragments of 125I-labelled vWF (21, 23). We have also reported on the RBD, CBD and the domain on vWF involved in the binding to thrombin activated platelets (thrombin binding domain, TBD) using vWF-fragments prepared by digestion with staphylococcal protease V8 (25). In the present study, we have digested 125I-vWF with TPCK-trypsin and we have performed at various times of digestion immuno-precipitation with Mab 9, the antibody inhibiting binding of vWF to thrombin activated platelets. The data were compared with the immunoprecipitation patterns simultaneously obtained with CLB-RAg 35 which inhibits binding of vWF in the presence of ristocetin and with CLB-RAg 201, which inhibits binding of vWF to collagen. At 90 min, Mab 9 and CLB-RAg 201 precipitated similar high molecular weight bands, whereas CLB-RAg 35 precipitated bands at 180 and 120 kDa. After 24 h, Mab 9 precipitated bands at 200, 155, 116 and 85 kDa; CLB-RAg 201 precipitated a band at 48 kDa and CLB-RAg 35 a band at 116 kDa. Two-dimensional electrophoresis demonstrated that the high molecular weight bands, precipitated by Mab 9 and CLB-RAg 201 at 90 min, were identical. The 116 kDa fragment recognized by CLB-RAg 35 had a different subunit composition than the 116 kDa fragment precipitated by Mab 9.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Comparison of tryptic fragments of von Willebrand factor involved in binding to thrombin-activated platelets with fragments involved in ristocetin-induced binding and binding to collagen. 355 Nov 82

A 34-year-old woman patient was found to have a chronic hereditary haemolytic anaemia. No abnormal haemoglobin band was detected by conventional electrophoresis, but a slow beta chain could be separated on urea-carboxymethyl cellulose chromatography. Investigations of the patient's haemoglobin revealed an unstable component. Analyses of chemical structure, including isolation and TPCK trypsin digestion of the abnormal globin chain. HPL chromatography, amino acid composition as well as sequence determination of the abnormal peptide, indicated that a glutamine was replaced by a proline at position beta 131 (H9). Biosynthesis studies demonstrated a normal rate of synthesis but relatively fast degradation of the mutant beta chain. The new variant is named as Hb Shanghai according to the place where it was discovered.
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PMID:A new unstable haemoglobin variant: Hb Shanghai [beta 131(H9)Gln----Pro] found in China. 367 9

A novel acetylating agent, methyl acetyl phosphate (MAP), has been designed to react with a nucleophile near an anion binding site of proteins. We examined the effect of MAP on hemoglobin (Hb), which has a well defined binding site for 2,3-diphosphoglycerate (DPG), to determine whether this reagent recognizes the DPG binding site. The progress of the reaction was monitored by ion-exchange high-performance liquid chromatography (HPLC) on a TSK CM-SW column. Modified Hb was initially chromatographed on CM-52 and then separated into its component chains. The alpha- and beta-chains from modified and unmodified Hb were digested by TPCK-trypsin. The peptide mixtures were chromatographed on Whatman ODS-3 reversed-phase HPLC columns and the peptide maps of modified and unmodified chains were compared. The peaks formed by the modification with MAP were further purified on YMC ODS-S5 columns and then subjected to amino acid analysis on a Dionex D-500 instrument after acid hydrolysis. We found that the newly formed peptides are beta T1 and beta T14 + 15 and that the loss of a peptide corresponding to beta T9 and beta T 10 + 11 is significant. No change in the alpha-chains was observed. The results suggest that MAP is indeed specific for the DPG binding site, as the above peptides contain the amino acid residues involved in the binding of DPG. We have assigned the acetylation sites as Val-1(beta), Lys-82(beta) and Lys-144(beta).
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PMID:Methyl acetyl phosphate: a novel acetylating agent. Its site-specific modification of human hemoglobin A. 373 26


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