Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0272170 (SDS)
 50,377 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. A neutral thiol protease was isolated from the extract of larvae of the mammalian trematode parasite, Paragonimus westermani metacercariae, by arginine-Sepharose, Ultrogel AcA-54 and DEAE-toyopearl column chromatography, measuring its activity by the hydrolysis of Boc-Val-Leu-Lys-MCA as a substrate. 2. The molecular weight of the purified enzyme was estimated to be 22,000 as a single polypeptide by SDS-polyacrylamide gel electrophoresis and was estimated to be 20,000 by size exclusion high-performance liquid chromatography. 3. The activity was suppressed by antipain, E-64, leupeptin, chymostatin, N-tosyl-L-lysine chloromethyl ketone, but was not affected by metallo protease inhibitors or serine protease inhibitors. 4. Studies on the substrate specificity showed that the enzyme hydrolyzed Boc-Val-Leu-Lys-MCA, Z-Phe-Arg-MCA, fluorescein isothiocyanate-labeled collagen, azocoll and casein. 5. The enzyme was found to hydrolyze peptide bonds of oxidized insulin B chain preferentially at the carboxy side of hydrophobic and basic amino acids.
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PMID:Purification and properties of a neutral thiol protease from larval trematode parasite Paragonimus westermani metacercariae. 330 26

Protease and antiprotease activities were estimated in nasal secretions from patients with chronic sinusitis and nasal allergy, using [3H]-casein as substrate. In the purulent nasal secretions, strong protease activity was measured, but there was less activity in the allergic nasal secretions. In contrast, trypsin inhibitory activity in allergic nasal secretions was much higher than in nasal secretions from the patients with chronic sinusitis. A protease inhibitor was partially isolated from nasal secretions of the nasal allergic patients by Sephadex G-150 gel chromatography and characterized. This protease inhibitor has an apparent molecular weight of 10,000 D, determined by SDS-polyacrylamidegel electrophoresis. It depresses the activities of bovine pancreatic trypsin, bovine pancreatic chymotrypsin and proteases in nasal purulent secretions, whereas it does not inhibit porcine pancreatic elastase, papain, or human plasmin.
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PMID:A protease inhibitor from human allergic nasal secretions. 332 29

Four distinct tyrosine protein kinases active on poly(Glu4,Tyr1) and angiotensin II, and operationally termed TPK-I, TPK-IIA, TPK-IIB and TPK-III have been resolved and partially purified from rat spleen particulate fraction by combining DEAE-Sepharose, heparin-Sepharose, phosphocellulose and polylysine-agarose chromatographies. Once partially purified all of them are free of Ser/Thr-specific protein kinase activity as judged using casein, histones, protamine and the peptide Arg-Arg-Ala-Ser-Val-Ala as substrates. TPK-I (apparent molecular mass 64 kDa, by gel filtration) and TPK-IIA (54 kDa) share several properties, including substrate specificity and stimulation by heparin; the latter however is much more responsive to polylysine then the former (10- and 3-fold maximum stimulation, respectively). Conversely TPK-IIB (51 kDa) is markedly inhibited by heparin and it is also characterized by its unique substrate specificity: unlike the other three tyrosine protein kinases it by far prefers the tetrapeptide Glu-Tyr-Ala-Ala over the decapeptide Asp-Ala-Glu-Tyr-Ala-Ala-Arg-Arg-Arg-Gly and readily phosphorylates band-3 protein of red cell membrane. The unusual preference for Mg2+ over Mn2+ as activator and the capability to phosphorylate calmodulin distinguish TPK-III (61 kDa) from the other isoenzymes. Moreover TPK-III is insensitive to heparin and polylysine and is inhibited by quercetin much more efficiently than the other enzymes (I50 = 10 microM). Upon incubation with [gamma-32P]ATP, TPK-I, TPK-IIA and TPK-III give rise to alkali-stable radiolabeled components of 61, 55 and 52 kDa respectively, as evaluated by PAGE/SDS. In every case such a radiolabeling takes place also in the presence of a large excess of phosphorylatable substrate (angiotensin II) while it is readily reversed by isotopic dilution with 10-fold excess unlabeled ATP, supporting the view that it represents an autophosphorylation process. No (auto)phosphorylation product(s) could be detected in TPK-IIB even if its amount, in terms of catalytic activity, was 10-fold higher than that of the others.
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PMID:Characterization of four tyrosine protein kinases from the particulate fraction of rat spleen. 335 7

A cytosolic protein-tyrosine kinase has been highly purified from bovine platelets using [Val5]angiotensin II as a substrate. The purification procedure involves sequential column chromatography on phosphocellulose, Sephacryl S-200, poly(L-lysine)-agarose, casein-Sepharose 4B and 2',5'-ADP-Sepharose 4B. Analysis of the most highly purified preparations by SDS/polyacrylamide gel electrophoresis revealed a major silver-stained band of molecular mass 71 kDa. This molecular mass was consistent with results obtained from sucrose density gradient centrifugation, indicating that the enzyme exists as a monomer. The purified kinase, called CPTK 71, efficiently phosphorylated tubulin and p36 (calpactin 1 heavy chain). However, it did not phosphorylate H1 histone. Half-maximal enzyme activity was observed at 2.2 microM ATP, and Mn2+, Co2+ and Mg2+ were effective divalent metal ions for the expression of activity. Insulin, epidermal growth factor, and platelet-derived growth factor had little or no effect on the kinase activity of CPTK 71. CPTK 71 had no immunological cross-reactivity with pp60src. These results suggest that CPTK 71 is a novel type of protein-tyrosine kinases among the enzymes so far reported.
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PMID:Purification and characterization of cytosolic protein-tyrosine kinase from bovine platelets. 339 Nov 64

In order to study the hormonal regulation of gene expression in mammary epithelial cells, we isolated a prolactin-responsive cell clone, HC11, from the COMMA-1D mouse mammary epithelial cell line. Clone HC11 was selected as a unique example of a cloned mouse mammary epithelial cell which has no requirement for complex, exogenously added, extracellular matrix or co-cultivation with other cell types for the prolactin-dependent in vitro induction of the endogenous beta-casein gene by lactogenic hormones. Induction of beta-casein mRNA is rapid and was detected 3 h after hormone stimulation. A prolactin-dependent increase in the rate of transcription of the beta-casein gene was shown in an in vitro nuclear transcription assay. beta-Casein protein was detected in an immunoblot assay after 24 h, and further accumulated during 5 days of hormone treatment. To identify low-abundance proteins induced directly after prolactin stimulation, mRNA was accumulated during 5 h of stimulation of HC11 cells with prolactin in the presence of cycloheximide. Following cycloheximide removal, the mRNA was translated into protein during a 60-min [35S]methionine pulse and the proteins were resolved by DEAE ion exchange HPLC and SDS-PAGE. A strong induction of a 120-kd cytosolic protein was detected which was maximally expressed within 6 h of hormone stimulation.
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PMID:Prolactin regulation of beta-casein gene expression and of a cytosolic 120-kd protein in a cloned mouse mammary epithelial cell line. 341 34

The proteinase secreted from Thermomonospora fusca YX grown on cellulose was purified by (NH4)2SO4 fractionation and cation-exchange chromatography. The isolated proteinase readily hydrolysed several proteins and demonstrated activity towards casein from 35 to 95 degrees C (at pH 8.0) with maximum activity at 80 degrees C. It exhibited broad pH and ionic-strength optima centered at pH 9.0 and 0.2 M-NaCl respectively, and it retained high activity in the presence of 2% (w/v) SDS, 20 mM-dithiothreitol and 1.0 M-NaCl. The proteinase, which was fully inhibited by phenylmethanesulphonyl fluoride, had an Mr of 14,500 and an isoelectric point at 9.21. A measurement of proteinase thermal stability demonstrated a T50% (15 min) of 85 degrees C at pH 4.5.
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PMID:Purification and characterization of the heat-stable serine proteinase from Thermomonospora fusca YX. 347 79

A 200-fold purification of the maturation-promoting factor or MPF from unfertilized eggs of Xenopus laevis is reported for the first time. Purification was achieved by three successive column chromatographies on hydroxyapatite, trisacryl blue and L-arginine-agarose. The presence of MPF was assessed by the usual maturation criteria after injections of test material into immature stage VI unstimulated X. laevis oocytes: the precocious appearance of the maturation spot (within 45-120 min), the germinal vesicle breakdown, the presence of the first polar body and the second metaphase spindle. Purification was monitored by the decrease of the minimal amount of protein injected in a constant volume (50 nl) required to induce 50% frequency of germinal vesicle breakdown. This amount decreased from 500 ng in the crude extract to 2.5 ng in the 200-fold purified material. Analysis by SDS-PAGE of the crude extract showed about 40 Coomassie-blue-stained polypeptides with molecular masses ranging from 300 kDa to 20 kDa, whereas in the 200-fold purified MPF only 5 stained polypeptides were revealed, with molecular masses of 62, 53, 49, 39 and 37 kDa. In vitro phosphorylations for the detection of kinase activities for endogenous and exogenous substrates were monitored by analysis of autoradiograms of SDS-PAGE, after treatment of fractions with [gamma-32P]ATP. Only inactive fractions eluted from columns ahead of MPF, and fractions containing MPF activity were tested. Phosphorylation of numerous stained polypeptides was demonstrated in the crude MPF extract and exogenous substrates such as phosvitin, casein and histone type II-AS were also strongly phosphorylated. In the MPF fraction, purified on hydroxyapatite, a polypeptide of 53 kDa was more highly and specifically phosphorylated and the presence of kinase activities was observed for the above three exogenous substrates. In the 100-fold and 200-fold purified MPF, phosphorylation of endogenous substrates could not be shown and kinase activities for the above three substrates were drastically decreased as compared with the crude and purified MPF obtained after hydroxyapatite column chromatography. However, neither endogenous phosphorylations nor kinase activities with the above exogenous substrates could be shown in inactive fractions eluted ahead of MPF at the different purification steps. Some characteristics of the purified material are also described in this paper.
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PMID:Partial purification of the maturation-promoting factor MPF from unfertilized eggs of Xenopus laevis. 353 98

Proteases were detected in aqueous extracts of Dirofilaria immitis microfilariae. Enzymes within the extract were capable of hydrolyzing Azocoll, a general protease substrate, at pH's 7, 8, and 9. Sensitivities to a variety of protease inhibitors indicated that multiple azocollytic enzymes were present in the extract, most prominent of which appear to belong to the serine class of proteases. By incorporating various substrates into the matrices of polyacrylamide gels, 2 SDS-resistant, mercaptoethanol-sensitive proteases in the MF extract were identified at 22 and 76 kDa. These proteases showed differential abilities to digest casein, fibrinogen, hemoglobin, and IgG. The MF extract hydrolyzed radiolabeled IgG into 8-10-kDa fragments following a 20-hr incubation. A similar degree of digestion was observed in 2 hr when viable microfilariae were used. The potential significance of these proteases in the evasion of host effector mechanisms is discussed.
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PMID:Proteolytic cleavage of IgG and other protein substrates by Dirofilaria immitis microfilarial enzymes. 355 16

Tropomyosin kinase is partially purified from 14-day-old chicken embryos using DEAE-cellulose, cellulose phosphate and gel filtration chromatography. The purest enzyme preparation consists of two major bands of Mr = 76,000 and 43,000 on SDS-polyacrylamide gel electrophoresis. The molecular weight of the enzyme is 250,000 determined by gel filtration chromatography. It phosphorylates casein and skeletal tropomyosin equally well but histone and phosvitin at a much slower rate. Smooth muscle myosin light chain, tropomyosin from platelet, erythrocyte and smooth muscle are not phosphorylated. The apparent Km for skeletal alpha-tropomyosin and ATP is 50 microM and 200 microM, respectively. Vmax varies between 100-300 nmol/min per mg depending on the purity of the preparation. Mg2+ and dithiothreitol are essential for activity but Ca+, calmodulin and cAMP are not required. The optimum temperature is 37 degrees C and optimum pH is about 7.5. Heparin, a potent inhibitor of casein kinase II, has no inhibitory effect on the enzyme. Similar tropomyosin kinase activity is not detected in skeletal muscle in adult rabbit and chicken. The tropomyosin kinase described here represents a hitherto uncharacterized kinase responsible for phosphorylation of tropomyosin in the chicken embryo.
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PMID:Isolation and characterization of tropomyosin kinase from chicken embryo. 359 68

1. The digestion of zinc present in human milk, cow's milk and a commercial babyfood was compared, using the piglet as a model for the human infant. 2. In piglets given human milk the pH of stomach contents was approximately 1 and 0.4 units lower than that of animals given respectively cow's milk and babyfood. The pH values of intestinal contents were approximately neutral and did not vary with the type of feed. 3. Hard casein curds were present throughout the stomachs and small intestines of animals fed on cow's milk or babyfood and between 55 and 70% Zn in these digesta samples were recovered in an insoluble form by centrifugation. In contrast, little solid material was observed in the digesta of animals fed on human milk, and 57 and 93% respectively of the Zn in digesta were recovered in a soluble form in the stomach and small intestine. 4. Soluble fractions prepared by centrifugation of digesta were analysed by filtration on Sephadex G-150. After any of the three feeds, soluble Zn in stomach contents was mainly in a low-molecular-weight form. In intestinal samples, however, Zn was present in low- and high-molecular-weight forms. Whilst there were similar amounts of Zn in the low-molecular-weight form in all samples, approximately three times as much of the total intestinal Zn was in a soluble high-molecular-weight form complexed to proteins in the animals fed on human milk compared with those fed on cow's milk or babyfood. 5. Analysis of protein-bound soluble Zn in intestinal samples on SDS-polyacrylamide gels resulted in a similar pattern of proteins for all feeds. Results indicated that at least some of these proteins were derived from intestinal secretions of the piglet. 6. Some implications of these results in respect of the mode of digestion of Zn and its biological availability to the human infant are discussed.
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PMID:Digestion of the zinc in human milk, cow's milk and a commercial babyfood: some implications for human infant nutrition. 367 53


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