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)

Several kinds of modified chymotrypsin were prepared with water-soluble acylating reagents, and their characteristics after hydrolyzing with unmodified chymotrypsin in aqueous-N,N'-dimethylformamide (DMF) media were compared. It was found that chymotrypsin (Csin), of which a 20% amino group was modified with a benzyloxycarbonyl group (Z(20)Csin), had more favorable characteristics than unmodified chymotrypsin with regard to hydrolytic activity in an aqueous DMF media. We also investigated the Z(20)Csin-catalyzed peptide synthesis in two different solution systems. In the one-layer system containing water and DMF, Z(20)Csin catalyzed the peptide bond formation in a higher yield than that by unmodifide chymotrypsin and enabled a synthetic reaction in even an 80% (v/v) DMF media, in which the hydrolytic reaction could not be carried out. Z(20)Csin catalyzed the condensation between some N-acyl amino acids or peptide derivatives and amino acids in 90% ethylacetate, 90% hexane or 50% benzene. This latter method employs a two-layer system, and the modified enzyme may be able to reduce the number of synthetic steps when preparing acyl peptides.
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PMID:Characteristics of chymotrypsin modified with water-soluble acylating reagents and its peptide synthesis ability in aqueous organic media. 136 28

Chymotrypsin (EC 3.4.21.1) powder suspended in hexane in the presence of Na2CO3.10H2O is a good catalyst for peptide synthesis. The salt hydrate releases water to fix the thermodynamic water activity of the system in accord with its dissociation pressure. Salt hydrates can be useful to buffer water activity in mainly organic enzyme reaction mixtures at a value permitting activity of the catalyst while minimising hydrolytic side reactions.
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PMID:Salt hydrates buffer water activity during chymotrypsin-catalysed peptide synthesis. 185 22

Acylation of protein amino groups by N'-hydroxysuccinimide ester of N-succinyl phosphatidylethanolamine in reverse micelles of diisooctylsulphosuccinate in hexane was studied. The experiment in a model system (glycine solution in reverse micelles) showed rate of acylation of amino groups to be by over an order of magnitude higher than rate of hydrolysis. Water-soluble proteins (alpha-chymotrypsin and IgG), modified by means of this method, can effectively bind liposomes without disturbing the integrity of liposomal membrane.
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PMID:[Covalent binding of phospholipids to protein globule in a system of reverse micelles]. 342 10

Rats pretreated with xylene or phenobarbital, and then exposed to n-hexane, exhibited a markedly increased peak serum concentration of the neurotoxic metabolite 2,5-hexanedione. In order to elucidate the mechanism underlying this synergistic effect, the major liver microsomal cytochrome P-450 isozymes induced by xylene and phenobarbital, respectively, were purified. In a reconstituted system both isozymes showed a high enzymatic activity with n-hexane as the substrate. Turnover numbers for the formation of 2-hexanol were 24 and 27 for the xylene- and phenobarbital-induced isozyme, respectively. The turnover numbers for 7-ethoxycoumarin, benzo[a]pyrene, and 1,1,2,2-tetrachloroethane were also in the same range for the two cytochrome P-450 preparations. The isozyme induced by xylene had an amino acid composition very similar to that of the phenobarbital-induced isozyme, and the purified proteins had identical electrophoretic mobilities on polyacrylamide gels in the presence of sodium dodecyl sulfate. Furthermore, similar peptide maps were obtained following digestion with alpha-chymotrypsin and papain, and each isozyme yielded a single immunoprecipitin band upon reaction with the immunoglobulin G fraction from rabbits immunized with the phenobarbital-induced enzyme. We conclude that xylene induces a rat liver microsomal cytochrome P-450 isozyme very similar to the major isozyme induced by phenobarbital and that this induction is the probable explanation for the enhanced formation of 2,5-hexanedione from n-hexane in vivo.
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PMID:Xylene induces a cytochrome P-450 isozyme in rat liver similar to the major isozyme induced by phenobarbital. 686 1

The presence of neutrophils within the lung is a characteristic feature of a variety of lung diseases. To evaluate the potential role of alveolar macrophages in modulating the migration of neutrophils to the lung, normal human alveolar macrophages obtained from volunteers by bronchopulmonary lavage, were exposed for various periods of time in vitro to heat-killed microorganisms, and noninfectious particulates, immune complexes, and the macrophage supernates were evaluated for chemotactic activity. The microorganisms, noninfectious particulates, and immune complexes were chosen as stimuli for alveolar macrophages because these stimuli are representative of a spectrum of pathogenic agents that cause neutrophil accumulation in the lower respiratory tract. After incubation with each of these stimuli, alveolar macrophages released low molecular weight (400-600) chemotactic factor(s) (alveolar macrophage-derived chemotactic factor[s] [AMCF]) with relatively more activity for neutrophils than monocytes or eosinophils. Checker-board analysis of the AMCF revealed that the factor was primarily chemotactic and not chemokinetic for neutrophils. The selectivity for neutrophils vs. monocytes could not be explained by a selective deactivation of monocytes, because the AMCF was more potent in deactivating neutrophils than monocytes. Partial characterization of AMCF demonstrated it was heterogeneous with the following features: (a) stable to heating at 56 and 100 degrees C for 30 min; (b) stable over a pH range of 1.0 to 12.0 for 60 min; (c) stable after exposure to trypsin, papain, chymotrypsin, collagenase, and elastase; (d) partially inhibited by serum chemotactic factor inhibitor(s); (e) two major isoelectric points (pI 7.6 and 5.2); and (f) partially extractable into ethyl acetate, ether, and hexane. Although AMCF was, at least, partially lipid in nature, it did not appear to be similar to previously described lipid chemotactic factors (e.g., hydroxy-derivatives of 5,8,10,14-eicosatetraenoic acid); analysis by gas chromatography-mass spectrophotometry of AMCF extracted into ethyl acetate did not reveal the presence of 5,8,10,14-eicosatetraenoic acid. The macrophage supernates containing the AMCF also stimulated normal human neutrophils to release lysozyme and lactoferrin but not lactate dehydrogenase. These studies suggest that a wide variety of potentially pathogenic stimuli induce normal alveolar macrophages to generate a low molecular weight chemotactic factor(s) that preferentially attracts neutrophils. Because alveolar macrophages are normal residents of alveoli, it is likely that by releasing this factor(s) macrophages play a significant role in amplifying the inflammatory processes seen in many acute and chronic lung diseases.
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PMID:Human alveolar macrophage-derived chemotactic factor for neutrophils. Stimuli and partial characterization. 699 85

A theoretical analysis of the causes of chemical equilibrium shift (i.e., change in product yield), accompanying replacement of water as the reaction medium by biphasic system "water-water-immiscible organic solvent", is given. A model is described, which is based on equilibrium partition of reagents between the two phases and establishment of a chemical equilibrium. In terms of this model, the apparent equilibrium constant, Kbiphasic which is evolved from the equilibrium concentrations of the reagents referred to the total volume of the biphasic system, should depend on the ratio of the volume of the organic and aqueous phases and the partition coefficients. The theoretical dependences were verified experimentally. Firstly, it was shown that for oxidation of isobutanol into isobutyraldehyde, catalyzed by alcohol dehydrogenase, the equilibrium shift that takes place in a water-hexane biphasic system is determined by the partition coefficients of the reagents found in an independent experiment. Varying the composition of the organic phase (hexane or ethyl acetate and their mixtures), the equilibrium could be shifted (compared to the aqueous solutions) towards both the initial reagents and the end-products: thereby changing the apparent equilibrium constant by two orders of magnitude. Secondly, alpha-chymotrypsin-catalyzed synthesis of ethyl ester of N-benzoyl-L-phenylalanine (from the respective acid and alcohol) in a biphasic system containing chloroform, benzene, carbon tetrachloride or diethyl ether, has been studied. The apparent equilibrium constant has been experimentally demonstrated to depend on the ratio of the aqueous and organic phase volumes in an extreme fashion, as has been predicted by the theory. The yield of the product in the reaction optimum amounts to 100%, whereas in water it is as low as 0.01%.
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PMID:Enzymatic synthesis in biphasic aqueous-organic systems. I. Chemical equilibrium shift. 701 6

Crystals of gamma-chymotrypsin grown in aqueous solution were soaked in n-hexane, and the structures of both the soaked and the native crystals were determined to 2.2-A resolution. Seven hexane molecules and 130 water molecules were found in the hexane-soaked crystals. Two of the seven hexane molecules are found near the active site, and the rest are close to hydrophobic regions on or near the surface of the enzyme. In the hexane structure, water molecules that were not observed in the native structure form a clathrate around one of the hexane molecules. Only 97 water molecules were found in the native structure. The temperature factors for atoms in the hexane environment are lower than those in the aqueous environment. There are significant changes between the two structures in the side chains of both polar and neutral residues, particularly in the vicinity of the hexane molecules. These changes have perturbed the hydrogen-bonding patterns. The electron density for the peptide bound in the active site has been dramatically altered in hexane and appears to be tetrahedral at the carbon that is covalently bound to Ser 195. The crystalline enzyme retains its active conformation in the nonpolar medium and can catalyze both hydrolysis and synthesis reactions in hexane.
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PMID:X-ray crystal structure of gamma-chymotrypsin in hexane. 800 97

Effects of membrane phospholipids on Na(+)-Ca2+ exchange and Na+ channel currents were studied in giant excised cardiac sarcolemmal patches. Phospholipids were suspended in an inert vehicle of alpha-tocopherol acetate and hexane and were then directly applied to the side of patch electrodes at a short distance from the tip during current recording. Phosphatidylserine strongly stimulated outward Na(+)-Ca2+ exchange current and altered the kinetics of cytoplasmic Na(+)- and Ca(2+)-dependent secondary modulation. This effect was partially reversed by phosphatidylcholine. Prolonged treatment with phosphatidylserine eliminated the inactivation transient normally observed upon rapid application of cytoplasmic Na+ but left cytoplasmic Ca2+ dependence largely intact. In such cases, subsequent chymotrypsin treatment removed cytoplasmic Ca2+ dependence, but had no further stimulatory effect, indicating maximum alleviation of inactivation by phosphatidylserine. While these results indicate that phosphatidylserine acts on a cytoplasmic, protease-sensitive regulatory domain of the exchanger, phosphatidylserine also stimulated the exchange current after deregulation by chymotrypsin, indicating an effect on the exchange mechanism itself. As in other myocyte preparations, cardiac Na+ currents in giant patches undergo a time-dependent negative shift in the voltage dependence of steady-state inactivation. Loss of phosphatidylserine from the cytoplasmic leaflet (i.e. loss of transbilayer asymmetry of phosphatidylserine distribution) does not appear to be the underlying cause, since phosphatidylserine did not reverse this shift, despite stimulation of Na(+)-Ca2+ exchange current in the same patches.
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PMID:A novel method for direct application of phospholipids to giant excised membrane patches in the study of sodium-calcium exchange and sodium channel currents. 839 64

Despite the extensive use and study of enzymes suspended in organic solvents, whether activity differences between different preparations can be accounted for by differences in protein secondary structure is still unknown. To address this issue, in the current study two model enzymes, alpha-chymotrypsin and subtilisin Carlsberg, were lyophilized and suspended in both polar and nonpolar organic solvents. The secondary structures of the proteins in the initial aqueous solution, in the lyophilized powder, and in the subsequent suspensions in organic solvents were determined using infrared spectroscopy. Lyophilization perturbed the secondary structure of both enzymes. With alpha-chymotrypsin, lyophilization from buffer followed by suspension in ethanol, hexane, or pyridine did not alter the unfolded structure observed in the dried powder. In contrast, with subtilisin Carlsberg, suspension of the dried enzyme in ethanol led to further perturbation of structure, whereas in hexane, and more so in pyridine, there was some return toward native structure. Lyophilization of the aqueous protein solutions in the presence of either trehalose or sorbitol led to retention of more native-like structure of both enzymes in the dried solid. However, large structural perturbations arose when these samples were suspended in organic solvents. The only exception was the subtilisin-trehalose mixture, which regained some native structure in ethanol and hexane. The greatest changes were noted in samples suspended in pyridine, in which the infrared spectra indicated extensive intermolecular beta-sheet formation from protein aggregates. There was not any consistent correlation between activity in organic solvents and either the initial structure obtained in the dried powders or the final structure when suspended in organic solvents. Nor could differences in residual water contents in dried samples or the total water content in the organic solvent reaction system account for the activity differences.
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PMID:Effect of secondary structure on the activity of enzymes suspended in organic solvents. 890 Apr 18

Five stains of Bifidobacterium bifidum (ATCC 11863 and 29591, and NCFB 1453, 1454, 1455) were examined for production of bacteriocins in MRS broth with 0.05% cysteine. Only strain NCFB 1454 excreted a bacteriocin into the broth: it was designated bifidocin B. Bifidocin B was sensitive to several proteolytic enzymes (protease IV, pronase E, protease XVII, proteinase K, trypsin, alpha-chymotrypsin, papain, and pepsin), but was resistant to catalase, peroxidase, lipase, lysozyme, cellulase, ribonuclease A, and amylases. It was also resistant to organic solvents such as ethyl alcohol, acetone, hexane, chloroform, methanol, and ether, and to heating at 90 degrees C for 15, 30, and 60 min or at 121 degrees C for 15 min. Bifidocin B remained active after storage at -20 or -7 degrees C for 3 months and retained biological activity after exposure to pH values of 2 to 10. Bifidocin B was active against some food-borne pathogens and food spoilage bacteria such as Listeria, Enterococcus, Bacillus, Lactobacillus, Leuconostoc, and Pediococcus species but was not active against the other gram-positive and gram-negative bacteria tested. Bifidocin B was produced during exponential phase, reaching a maximum activity of 3,200 AU/ml at early stationary phase. Bifidocin B had a molecular mass of about 3.3 kDa as analyzed by Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
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PMID:Characterization and antimicrobial spectrum of bifidocin B, a bacteriocin produced by Bifidobacterium bifidum NCFB 1454. 970 52


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