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 antibiotic-producing bacterium repeatedly isolated from human feces was characterized by standard bacteriological methods. The bacterium is a gram-positive bacillus possessing morphological and physiological features similar to those of Bacillus subtilis, except that it lacks temperature-resistant spore formation and has peritrichous flagella. The cell-free antibiotic produced by the organism in vitro was effective against some gram-negative and gram-positive bacteria and yeast in minimum inhibitory concentration and antibiotic disk assays. The 1,200-dalton antibiotic had an optimal activity against Proteus vulgaris within the pH range of 5.7 to 6.8. The activity was totally destroyed by digestion with pronase and trypsin but was resistant to pepsin, chymotrypsin, papain, and nuclease digestion. In addition, the antibiotic activity against P. vulgaris was stable between pH 3 to 9 and within the temperature range of 20 to 100 degrees C when tested in the fermentation medium. The activity was only partially retained by membrane filters which normally retain globular proteins of molecular weights between 500 to 10,000. Electrophoresis in phosphate buffer indicated that the activity against P. vulgaris had an isoelectric point of approximately 6.45. These properties are compatible with the antibiotic activity associated with a small peptide.
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PMID:Identification of an antibiotic-producing bacterium from the human intestinal tract and characterization of its antimicrobial product. 62 95

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to examine the polypeptide patterns of rat liver rough and smooth endoplasmic reticulum (ER) membrane fractions stripped of ribosomes. Approximately 67 polypeptides were resolved from the rough ER membrane fraction. The polypeptide pattern of the smooth ER membrane fraction was similar to that of the rough ER membrane fraction, but exhibited substantially lower amounts of some seven polypeptides. Three of these polypeptides, of apparent molecular weights 63,000, 65,000, and 87,000, were of particular interest, as they could not be ascribed to contamination of stripped rough ER membrane fractions by residual ribosomal polypeptides. Conditions of treatment with low concentrations of trypsin were established that markedly diminished the capacity of the stripped rough ER membrane fraction to bind ribosomes in vitro and that also effected a partial detachment of ribosomes from nonstripped rough ER membranes; the results of electrophoretic analyses of rough ER membrane fractions treated in these manners are described. Comparison of the polypeptide patterns of guinea pig, mouse, and rabbit liver ER membrane fractions with rat liver ER membrane fractions revealed considerable variations in the distribution of the polypeptides of 63,000, 65,000, and 87,000 molecular weight among the ER membrane fractions of these species. The combined results of these studies indicate that the polypeptide of 87,000 molecular weight, although particularly sensitive to attack by trypsin, is not involved in the binding of ribosomes to the rough ER membrane fraction. Studies by others (cf. Kreibich, G., Grebenau, R., Mok, W., Pereyra, B., Rodriguez-Boulan, E., and Sabatini, D. D. (1977) Fed. Proc. 36, 656) have implicated the polypeptides of 63,000 and 65,000 molecular weight in this process. The patterns of phosphorylated polypeptides of rough and smooth ER membrane fractions of rat and mouse liver were also examined, using labeling in vivo with sodium [32p]phosphate or in vitro with [gamma-32P]ATP. Approximately 25 phosphorylated components were resolved by electrophoresis in the ER membrane fractions of both species. Evidence is presented that suggests that the great majority of these components are phosphopolypeptides. Differences were noted in the patterns of phosphorylation produced by in vivo and in vitro labeling; minor differences were also observed between the patterns of phosphorylation of the rough and smooth ER membrane fractions in either situation. The overall results afford an indirect approach toward evaluating the possible involvement of specific rough ER membrane polypeptides in ribosome-binding and reveal that liver ER membranes contain a substantially greater number of phosphorylated polypeptides thatn previously reported.
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PMID:Electrophoretic studies on liver endoplasmic reticulum membrane polypeptides and on their phosphorylation in vivo and in vitro. 63 54

The amino acid sequence around the pyridoxal 5'-phosphate binding site in potato phosphorylase was determined in order to compare it with those in phosphorylases from other sources having different regulatory properties. The potato enzyme was reduced by NaBH4 in the presence of urea, carboxymethylated, and digested with chymotrypsin and trypsin. Pyridoxyl peptides were isolated by the differential procedure using paper electrophoresis or DEAE-cellulose column chromatography. In Edman degradation of these peptides, pyridoxyllysine was identified as the phenylthiohydantoin derivative of pyridoxyllysine using a combination of thin-layer chromatography and the Pauli reaction. The sequence around pyridoxyllysine, comprising 57 amino acid residues, was determined except for a region with 6 amino acid residues. The pyridoxal 5'-phosphate binding site in potato phosphorylase showed a high homology with those of the rabbit muscle and yeast enzymes. This finding suggests that the cofactor should be directly related to the essential process of phosphorylase action.
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PMID:Amino acid sequence around the pyridoxal 5'-phosphate binding site in potato phosphorylase. 65 83

The sequence analysis of llama (Lama glama, Camelidae) hemoglobin is described. The chains were separated, cleaved by trypsin as previously described, quantitatively characterized and sequenced in the sequenator. The llama hemoglobin differs from the human hemoglobin in that it has 25 different amino acids in the alpha chain and 24 different amino acids in the beta chain. The interaction between protein and phosphate is discussed. The earlier finding that the O2 affinity of the llama hemoglobin is dependent on its content of 2, 3-bisphosphoglycerate is interpreted here as a mutation of the 2, 3-bisphosphoglycerate contact position beta2 His in human hemoglobin to beta2 Asn in llama hemoglobin, whereby one of the four 2, 3-bisphosphoglycerate contact points is interrupted. This interruption gives rise to a diminished reduction of intrinsic oxygen affinity in the hemoglobin molecule and explains, on a molecular basis, the increased oxygen affinity of the llama hemoglobin, and consequently, the high-altitude respiration of the llama. By analogy, the increased O2 affinity of human fetal hemoglobin is interpreted according to previous physiological investigations on blood and fetal hemoglobin by the inactivation of the phosphoglycerate contact point beta143 His in the adult hemoglobin by mutation to gamma 143 Ser in the fetal hemoglobin. With respect to respiration in horses (2, 3-bisphosphoglycerate contact beta2 Gln), measurement of atomic parameters show that the amido group of the glutamine is situated close enough to the 2, 3-bisphosphoglycerate oxygen to build a hydrogen bond with the phosphate. Consequently, the explanation of the low-altitude respiration of the horse lies in the fact that glutamine and histidine fulfill sterochemically an identical function.
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PMID:[The interaction between phosphate and protein, and the respiration of the llama, the human fetus and the horse (author's transl)]. 66 74

A method for the enrichment of live thyrotrophic pituitary cells is described. Pituitary glands of young male rats were removed into Earle's solution and dispersed in a 0.1% trypsin solution containing 0.5% bovine serum albumin, pH 7.2. Nylon fibres (25 microgram) were used for the separation of the thyrotrophic cells, by stringing them across a plastic frame which fitted a plastic Petri dish containing the cell suspension. The fibres were washed with light petroleum (b.p. 60--80 degree C) and carbon tetrachloride, hydrolysed with 3 M-HCL for 30 min at room temperature and washed with distilled water and phosphate-buffered saline (pH 7.2). The fibres were treated with thyrotrophin releasing hormone (TRH) alone or in the presence of soluble carbodiimide solution. After incubation for 1 h at room temperature, the fibres were transferred to a new Earle's medium and cells were released from the fibres by plucking them with a needle. The separated thyrotrophic cells were identified by radioimmunoassay and by electron microscopy. Using the above-mentioned methods, enrichment of thyrotrophic cells was obtained. Thus, the amounts of TSH, prolactin, LH and GH released, during 2 h of incubation, by 1.5 x 10(6) unseparated cells were 6.8 +/- 0.65, 4.1 +/- 0.47, 4.8 +/- 0.52 and 5.2 +/- 0.68 microgram respectively, while the same number of purified thyrotrophic cells released 76.1 +/- 0.42, 1.2 +/- 0.3, 0.6 +/- 0.35 and 1.6 +/- 0.22 microgram of the same hormones (means +/- S.E.M.).
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PMID:Separation of rat pituitary thyrotrophic cells. 68 67

Rabbit skeletal muscle glycogen phosphorylase b was covalently bound to oyster glycogen by means of cyanogen bromide. Removal of the unbound enzyme was achieved, using DEAE-Sephadex A-50 chromatography. Glycogen-bound phosphorylase b showed a higher affinity toward glucose 1-phosphate but a lower homotropic cooperativity, with respect to AMP activation, than the native enzyme. However, at low AMP concentrations conjugated phosphorylase b was as efficient as the free enzyme. It is of interest that glycogen-bound phosphorylase b exhibited catalytic activity upon its polysaccharide carrier. Kinetics of heat and cold inactivation indicated that the bound enzyme was considerably more resistant toward heat inactivation but less stable upon exposure to cold. It was shown also that both conjugated and native enzymes had identical pH optima, similar activity/temperature dependencies and the same resistance against trypsin inactivation.
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PMID:Phosphorylase b covalently bound to glycogen: properties of the complex. 68 38

When pancreatic DNase I is used as a specific biochemical reagent in the preparation of nuclear ribonucleic acids or nuclear proteins, freedom from contaminating ribonucleases or proteases is an important property of the enzyme preparation. A simple one-step procedure has been developed to effect complete removal of trypsin, chymotrypsin, and chymotrypsinogen by a combination of affinity chromatography and salting-out adsorption on lima bean protease inhibitor coupled to Sepharose (a column (0.9 X 60 cm) operated in series with a regeneratable 1-ml bed). Commercial preparations of DNase (about 10 mg) give a quantitative yield of the enzyme that is protease-free as evidenced by full stability for more than 10 days at pH 8 and 37 degrees C even in the absence of the protecting action of Ca2+. Removal of the last traces of RNase has been accomplished by affinity chromatography on a column (0.4 X 72 cm) of 5'-(4-aminophenyl-phosphoryl)-uridine 2'(3')-phosphate-Sepharose; the product is a highly active DNase that gives no detectable hydrolysis of RNA by assay on radioactive substrates.
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PMID:Preparation of protease-free and ribonuclease-free pancreatic deoxyribonuclease. 70 Dec 44

Limited digestion, with trypsin, of the fatty acid synthetase from rat mammary gland releases an enzymically active thioesterase component that, under denaturing conditions, consists of two major species of mol.wts. 35000 and 17500 and a minor species, mol.wt. 15,000. The 17500- and 150000-mol.wt. species are shown to originate from the 35000-mol.wt. species as a result of nicking by trypsin. The nicked polypeptides are enzymically active. The fatty acid synthetase is inhibited by [1,3-14C]di-isopropyl phosphorofluoridate, which is shown to bind to, and inactivate, two thioesterase active sites. When the [1,3-14C]di-isopropyl phosphate-labelled fatty acid synthetase is subjected to limited digestion with trypsin, all of the radioactivity is recovered in the isolated thioesterase component, i.e. in the 35000-mol.wt. polypeptide and its nicked products. Since the isolated thioesterase is shown to bind only one di-isopropyl phosphate residue per 35000-mol.wt. polypeptide, we conclude that the fatty acid synthetase has two thioesterase domains, both of which are removed by limited trypsin treatment.
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PMID:Release of two thioesterase domains from fatty acid synthetase by limited digestion with trypsin. 73 93

Critical mixtures of aqueous solutions of polymers can be separated into two or more immiscible phases. Particulate materials are distributed in such phase systems generally between one bulk phase and the interface between bulk phases. The distribution is described by a simple partition law and is quantitatively determined by temperature, interfacial tensions, the electromotic potential difference between phases and the nature of the particle surface. The effects on transfacial potential differences of varying polymer, NaCl and sodium phosphate concentrations in dextran/polyethylene glycol systems were studied: increase of polyethylene glycol concentration increased the potential; addition of up to 40 mM NaCl progressively reduced the potential to zero: very small (less than 10 mM) concentrations of sodium phosphate buffer increased the potential, but further increase caused a fall in potential, which was less marked than that caused by equivalent concentrations of NaCl. The partition properties of a variety of cells, native or modified by treatment with trypsin, neuraminidase or maleic anhydride, were studied. In systems containing greater than 40 mM NaCl no difference in partition patterns for modifications of each cell type was observed. In systems containing no NaCl the partition pattern was highly dependent on the nature of the modification. From the behaviour of such models, polymer-electrolyte phase systems suitable for study of cell surface modification involving change, or no change, in net surface have been identified.
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PMID:Cell partition: a study of parameters affecting the partition phenomenon. 76 Aug 20

Native spectrin has trypsin-susceptible sites spaced at a constant molecular weight interval. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of spectrin treated with trypsin at low salt concentrations shows a ladder of fragments spaced at approximately 8,000-dalton intervals, from the intact Band 1 (240,000 daltons) and Band 2 (220,000 daltons) down to about 150,000 daltons. The five largest fragments were identified as products of Band 2 using tryptic 125I-peptide mapping of protein from gel slices. Endogenously incorporated [32P]phosphate is absent from the largest fragment, indicating that all phosphorylation sites on spectrin are within 8,000 daltons of a terminal of Band 2. Mapping of both [14C]carboxyamidomethylated cysteine-containing tryptic peptides and 125I-peptides reveals extensive sequence homology between the spectrin subunits. Further, only somewhat over half of the distinct spots expected from the cysteine content are found in both Band 1 and Band 2 peptides. These and the tryptic susceptibility results are interpretable as evidence for a repeating structure in spectrin.
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PMID:Structural studies on human spectrin. Comparison of subunits and fragmentation of native spectrin. 76 4


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