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:3.4.21.1 (chymotrypsin)
10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The trifunctional beta-subunit of anthranilate synthase complex of Neurospora crassa has been purified from a mutant which produces no detectable alpha-subunit. The isolated beta-subunit appeared to be a highly asymmetric dimer with a s20,w of 7.35 and an apparent molecular weight of 200,000 as determined by gel filtration on Sephacryl S-300 compared with a monomer molecular weight of approximately 84,000 Da as determined by sodium dodecyl sulfate-gel electrophoresis. The purified subunit was cleaved by elastase, trypsin, or chymotrypsin into fragments which retained the three enzyme activities. After elastase digestion, two active fragments were separated by gel filtration and ion exchange chromatography. A 30,000-Da fragment, which behaved as a monomer on gel filtration, interacted with free alpha-subunit to produce glutamine-dependent anthranilate synthase activity. A second 56,000-Da fragment, which behaved as an asymmetric dimer (apparent molecular weight 140,000) on gel filtration, retained both N-(5'-phosphoribosyl)anthranilate isomerase and indole-3-glycerol phosphate synthase activity. The failure to detect an NH2-terminal amino acid residue on either the intact beta-subunit or the 30,000-Da complementing fragment, while the 56,000-Da fragment possessed an NH2-terminal histidine residue, indicated that the complementing fragment was derived from the NH2-terminal sequence of the beta-subunit.
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PMID:Purification and characterization of the trifunctional beta-subunit of anthranilate synthase from Neurospora crassa. 621 92

Conditions have been found for limited proteolysis of purified tubulin, in which 70-90% of the molecules are cleaved at one or two sites. Thermolysin and chymotrypsin cleave the alpha and beta subunits, respectively, at single sites. Trypsin cleaves the alpha subunit at two sites. The chymotrypsin site and one of the trypsin sites are apparently inaccessible on assembled microtubules. The different samples of proteolyzed tubulin were all fully competent to assemble in a buffer containing 1 M sodium glutamate. In another buffer (50 mM morpholinoethanesulfonic acid, 3.4 M glycerol) tubulin digested by thermolysin assembled as well as native tubulin, but samples digested by chymotrypsin or trypsin would not assemble even at high protein concentrations.
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PMID:Assembly of proteolytically cleaved tubulin. 633 34

Calcium transport into everted membrane vesicles of Escherichia coli was found to have two components, one phosphate-dependent and the other phosphate-independent. In vesicles prepared in a glycerol buffer, calcium/proton exchange was phosphate independent but net uptake of 45Ca2+ required phosphate. In vesicles prepared in a sucrose buffer both phosphate-independent and phosphate-dependent accumulation of 45Ca2+ occurred. Both calcium/proton exchange and phosphate-independent uptake of 45Ca2+ were inactivated by treatment with trypsin, chymotrypsin, or N,N'-dicyclohexylcarbodiimide but not by N-ethylmaleimide. Phosphate-dependent uptake of 45Ca2+ was inhibited by N-ethylmaleimide but not by trypsin, chymotrypsin, or N,N'-dicyclohexylcarbodiimide. Under conditions permitting only phosphate-dependent uptake of 45Ca2+, concomitant uptake of 32Pi was also observed. Both uptake and efflux of the two ions occurred with a 1:1 ratio. These results imply the existence of two calcium transport systems in everted membrane vesicles, one of which catalyzes exchange of calcium ions and protons, the other of which catalyzes cotransport of calcium and phosphate.
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PMID:Calcium efflux from Escherichia coli. Evidence for two systems. 637 51

The effects of vegetarian fasting were evaluated in 14 grossly obese patients who participated in a program comprising 5 weeks' fasting in a lactovegetarian health center. Before and after the fasting period the patients were hospitalized and put on a standardized weight-maintaining diet; at the health center they consumed vegetable juices containing less than 1 MJ and 3 g of protein per day. The weight reduction (mean +/- S.D.) was 13.4 +/- 5.0 kg (from 132.0 +/- 27.2 to 118.6 +/- 16.1 kg). Except for the first few days the patients had no severe hunger sensations. No severe adverse clinical effects were noted. The laboratory status--comprising serum or plasma levels of minerals, protein, and lipids; hematological data; and variables reflecting liver and thyroid function--revealed abnormal group mean values only for ferritin and the acute-phase reactants haptoglobin, C-reactive protein, and anti-chymotrypsin in the obese. The levels of potassium, retinol-binding protein, and haptoglobin decreased, and aminotransferase and lactate dehydrogenase activities and free fatty acid and glycerol concentrations increased as a result of the fasting. The most striking effect of the weight reduction was an increase in the HDL cholesterol levels. Fasting according to the described regimen thus seems to provide a safe method for treatment of obese patients.
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PMID:Vegetarian fasting of obese patients: a clinical and biochemical evaluation. 713 69

The ultramicro (1-microL samples) assay of serum or plasma triglycerides that we describe here is potentially applicable to 1-nL samples and to isolated cells. This technically simple method involves only three reactions: (a) enzymic hydrolysis with lipase and alpha-chymotrypsin; (b) conversion by glycerol kinase of the liberated glycerol and of adenosine triphosphate added in excess to glycerol-1-phosphate and to adenosine diphosphate; and (c) assay of the residual adenosine triphosphate by the luciferin-luciferase reaction. The assay was optimized with respect to glycerol kinase, buffer, pH, temperature, and adenosine triphosphate. Performance characteristics compare well with those of traditional triglyceride assays.
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PMID:Ultramicro determination of serum triglycerides by bioluminescent assay. 746 Feb 77

Cucumisin (EC 3.4.21.25), a serine endopeptidase, was isolated by a simple purification procedure from the prince melon (Cucumis melo ssp. melo, cv. 'Prince Melon'). The enzyme is stable over a wide pH range (4-11) and to heat, 80% of its initial activity remaining even at pH 11.1 and at 60 degrees C for 20 min. The enzyme was inactive at 72 degrees C and pH 8.0, but 38% of the activity remained in the presence of 10% (w/v) glycerol. Caseinolysis by cucumisin indicated full activity in 8 M urea at pH 9.1 and 50 degrees C. Cucumisin was inactivated by treatment with trypsin at 37 degrees C for 24 h, but was not affected by alpha-chymotrypsin. The synthetic substrates benzyloxycarbonyltyrosine nitrophenyl ester (Z-Tyr-ONp) and benzoyltyrosine ethyl ester (Bz-Tyr-OEt) were cleaved, but Z-Lys-ONp and tosylarginine methyl ester (Tos-Arg-OMe) were not cleaved by cucumisin. Oxidized insulin B-chain was hydrolysed by cucumisin at 37 degrees C for 24 h, 21 cleavage sites being detected. Cucumisin could not cleave the C-termini of all the valine residues in the oxidized insulin B-chain molecule.
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PMID:Improved isolation, stability and substrate specificity of cucumisin, a plant serine endopeptidase. 757 59

The influence of the synthetic substrate (N-acetyl-L-tyrosine ethyl ester) and the different polyols (ethylene glycol, glycerol, erythritol, xylitol and sorbitol) on the thermostability of alpha-chymotrypsin at 60 degrees C have been studied. The results obtained showed an important stabilizing effect in the presence of both additives. In order to describe the kinetics of enzyme stabilization, the experimental results were analyzed by a four-parameters deactivation model with excellent agreement. In all cases, alpha-chymotrypsin exhibited non-first-order deactivation kinetics, corresponding to a two-step unimolecular mechanism, where the main protective effect of polyols was observed in the first-step of the deactivation profile. Thus, the presence of polyols increased the level of activity stabilization (alpha 1), and decreased the first-order deactivation rate constant (k1). Additionally, the experimental results were analyzed as a function of both, the change in the standard free energy of denaturation (delta(delta Gzero)), and a protective effect, defined as the ratio of alpha-chymotrypsin half-lives (with and without polyols), showing in both cases a clear stabilizing effect of these polyhydroxylic cosolvents for the enzyme. The overall protective effect of polyols was also simultaneously related to their concentration and their water-activity depressing power.
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PMID:Effect of polyols on alpha-chymotrypsin thermostability: a mechanistic analysis of the enzyme stabilization. 776 28

A diglyceride derivative of a pentapeptide renin inhibitor, the 1,3-dipalmitoyl-[Iva-Phe-Nle-Sta-Ala-Sta-acetyl]-glycerol was synthesized and tested in vitro as a potential prodrug for oral administration. The ability of the diglyceride analog to inhibit the renin activity was equivalent to that of the parent peptide after predigestion with pancreatic lipase. Furthermore, the presence of the palmitoyl groups was found to induce, in vitro, an efficient protection of the peptide from gastric and intestinal hydrolysis. During incubation with intestinal and gastric fluids, and with alpha-chymotrypsin and pancreatic lipase, the glycerolipidic derivative was more stable than the peptide alone. These results support the use of glycerolipidic prodrug for oral administration of peptides.
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PMID:Synthesis and in vitro study of a diglyceride prodrug of a peptide. 797 5

A detailed comparison has been made between dichroic steady-state spectroscopic properties at 77 K of several trimeric and monomeric forms of the major chlorophyll a/b binding protein (LHC-II) from pea. Monomeric forms were obtained by applying high concentrations of nonionic detergents, by a lipase treatment, or by a chymotrypsin/trypsin treatment. The latter treatments removed phosphatidyl glycerol essential for trimer formation. The absorption and dichroism spectra indicate that for trimeric LHC-II the chlorophyll b absorption region is centered around 649 nm and is composed of at least five subbands near 640, 647, 649, 652, and 656 nm. The chlorophyll a absorption region is centered around 670 nm and is composed of at least five bands near 661, 668, 671, 673, and 676 nm. The chlorophyll b band near 647 and 652 nm and the chlorophyll a bands near 668 and 673 nm are absent in the circular dichroism spectrum after monomerization. A configuration in which pigments of the same nature located on different monomers become excitonically coupled in the trimer could explain these results. In monomers obtained in high concentrations of nonionic detergents, no additional bands have disappeared, but the absorption spectra of the other two types of monomers lack the bands at 640 and 661 nm. These monomers have lost some chlorophyll a and b according to the fluorescence emission spectra, which show contributions from free chlorophyll a and b. The results suggest that phosphatidyl glycerol not only is involved in trimer formation but also has a structural role within the monomers.
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PMID:Spectroscopic characterization of three different monomeric forms of the main chlorophyll a/b binding protein from chloroplast membranes. 799 6

Following synthesis in the cytoplasm, the transforming proteins encoded by the retroviral oncogenes src, yes, fps, fes, and fgr form complexes with hsp90 and the hsp90 cohort p50. These cytoplasmic complexes are intermediates in the production of the mature membrane-associated kinase. However, soluble complexes between the nascent cellular homologs of these proteins and hsp90-p50 have not been readily detected [Brugge, J.S. (1986) Curr. Top. Microbiol. Immunol. 123, 1-22 and references therein]. In this paper, we have utilized protein synthesis in reticulocyte lysate to determine whether three cellular members of the src family of tyrosine kinases, myeloid-specific p59fgr, B cell-specific p59fgr, and p56lck, form complexes with hsp90. Following their synthesis, fast- and slow-sedimenting forms of these proteins can be separated on glycerol gradients. Anti-hsp90 monoclonal antibodies co-immunoadsorb the fast-sedimenting, but not the slow-sedimenting, forms of these kinases from gradient fractions. These hsp90 complexes can be detected in the complete absence of detergent. Conversely, an unrelated protein, firefly luciferase, does not form stable complexes with hsp90 following synthesis in reticulocyte lysate. Anti-p56lck antibodies specifically co-immunoadsorb hsp90 from protein synthesis reactions programmed with lckRNA. The fast-sedimenting, complex-bound form of p56lck is deficient in autophosphorylation activity and phosphorylates an exogenous substrate, acid-treated enolase, less efficiently than does the monomeric form. Fast-sedimenting p56lck is hypersentitive to limited proteolysis by chymotrypsin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Association of Hsp90 with cellular Src-family kinases in a cell-free system correlates with altered kinase structure and function. 804 79


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