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141.
During the last decade increasing attention has been given to the role of free radicals in biological oxidations. The subject has been of increasing interest to both the food scientist and the physiologist. Free radical scavengers in the form of both indigenous and added antioxidants are necessary for the successful preservation of food; free radicals are increasingly being implicated in the onset of, among others, ischaemic heart disease and for protection against these diseases it is suggested that the dietary intake of the antioxidant vitamins should be increased especially for diets high in polyunsaturated fats. 1,2 Convenience and snack foods which absorb substantial amounts of frying oils are being increasingly consumed. Since poly-unsaturated fatty acids are particularly susceptible to oxidation by free radicals during the storage, cooking and frying of foods, the potential risk of exposure to lipid degradation products' is likely to have increased. In foods the non-enzymic and lipoxygenase catalysed oxidation of polyunsaturated fatty acids, β-carotene and vitamin A can result in the loss of essential nutrients and the development of off-flavours. 相似文献
142.
Subtilisin Carlsberg was covalently attached to five macroporous acrylic supports of varying aquaphilicity (a measure of hydrophilicity). Kinetic parameters of the transesterification of S and R enantiomers of secphenethyl alcohol with vinyl butyrate, catalyzed by various immobilized subtilisins, were determined in anhydrous dioxane and acetonitrile. Enzyme enantioselectivity in acetonitrile, but not in dioxane, correlated with the aquaphilicity of the support; a mechanistic rationale for this phenomenon was proposed. Although the catalytic activity of immobilized subtilisin in anhydrous solvents strongly depended on enzyme pretreatment, the enantioselectivity was essential conserved. (c) 1994 John Wiley & Sons, Inc. 相似文献
143.
144.
Invertase as well as as amyloglucosidase were immobilized within asymmetyric ultrafiltration membranes that were prepared from polysulfone or homogeneously modified polysulfone. The chemical modification was carried out by sulfonation and halomethylation. This additional change of the surface properties of the capillaries within the membrane offers the possibilities for various types of enzyme fixation, namely adsorption, charge interactions, or covalent bonding. By variation of the immobilization conditions the distribution of the enzyme could be adjusted over the membrane's cross section. At a distinct enzyme concentration in the loading solution a homogeneous enzyme distribution within the membrane could be verified. This was shown by diffusion experiments. Under ultrafiltration conditions using a solution that contains membrane-impermeable macromolecules as well as a membrane-permeable solute like saccharose the residence time within the membrane was increased due to gel formation atop the membrane yet the kinetic was no affected. The nonpermeable soluble starch was not reacted by the amyloglucosidase membrane, indicating that the skin layer was free of enzymes. (c) 1994 John Wiley & Sons, Inc. 相似文献
145.
Specific quantifications of the major cellulolytic components of the Trichoderma reesei enzyme complex, i.e., endoglucanases I and III and cellobiohydrolases I and II, are described and, employing a defined mixture of these four cellulases reconstituted according to the composition of the native Trichoderma cellulase complex, used to determine the binding of each individual component onto filter paper. During substrate degradation by this enzyme mixture, the specific adsorption of each individual cellulase gradually increases and no preferential binding of one enzyme component in any particular phase of cellulose hydrolysis is found. T. reesei cellobiohydrolases I and II admixed with endoglucanases I and III represent a "full-value" cellulase system that is capable of degrading semicrystalline cellulose efficiently. In comparison with the crude Trichoderma enzyme complex, almost identical adsorption properties and similar hydrolytic efficiency are found for the reconstituted mixture. (c) 1994 John Wiley & Sons, Inc. 相似文献
146.
Nagayasu T Miyanaga M Tanaka T Sakiyama T Nakanishi K 《Biotechnology and bioengineering》1994,43(11):1108-1117
N-(Benzyloxycarbonyl)-L-aspartyl-L-phenylalanine methyl ester (Z-AspPheOMe), a precursor of the aspartame, and N-(benzyloxycarbonyl)-L-phenylalanyl-Lphenylalanine methyl ester (Z-PhePheOMe) were synthesized from the respective amino acid derivatives with an immobilized thermolysin (EC 3.4.24.4) in ethyl acetate. Various factors affecting the synthesis of these dipeptide precursors were clarified. The initial synthetic rate was the highest at the water content of 3.5% for both reactions. The substrate concentration dependencies of the initial synthetic rate of Z-AspkPheOMe and Z-PhePheOMe with the immobilized enzyme in ethyl acetate were different from those in an aqueous buffer solution saturated with ethyl acetate but similar to those in the aqueous/organic biphasic system using the free enzyme. Particularly, the initial synthetic rate of Z-AspPhOMe increased in order higher than first order with respect to the concentration of L-phenylalanine methyl ester (PheOMe), whereas it decreased sharply with the concentration of N-(benzyloxycarbonyl)-L-aspartic acid (Z-Asp). Such kinetic behavior could be explained by regarding the inside of the immobilized enzyme as being a biphasic mode composed from the organic phase and aqueous phase where the enzymatic reaction takes place. The reaction in the aqueous/organic biphasic system using the free enzyme could be simulated by taking into consideration the partition of the substrate and the initial rate of synthesis in the aqueous buffer saturated with ethyl acetate. Based on this analysis, the rate of reaction with the immobilized enzyme in ethyl acetate could also be predicted. Z-AsPheOMe and Z-PhePheOMe were synthesized by the fed-batch method where the acid component of the substrate was intermittently added during the course of reaction and by the batch method. In the synthesis of Z-AspPheOMe, the synthetic rate and maximum yield of reaction as well as the stability of the immobilized enzyme were higher in the fed-batch reaction than those in the batch reaction. In the synthesis of Z-PhePheOMe, the results obtained by both methods were similar. (c) 1994 John Wiley & Sons, Inc. 相似文献
147.
Plant defence systems induced by ozone 总被引:29,自引:9,他引:20
J. KANGASJÄRVI J. TALVINEN M. UTRIAINEN R. KARJALAINEN 《Plant, cell & environment》1994,17(7):783-794
Recent advances in the understanding of the molecular basis of plant response to ozone attack are reviewed. Plants grown in elevated atmospheric ozone are known to undergo several biochemical changes before any actual damage can be detected. These reactions include increases in the activities of enzymes associated with general plant defence mechanisms. Ozone exposure often causes a surge in the production of the plant hormone ethylene, as well as changes in polyamine metabolism and increases in the activities of several phenylpropanoid and flavonoid pathway enzymes. The activities of superoxide dismutase and peroxidases that protect cells from the oxidative damage caused by hydroxyl radicals, H2O2 and superoxides also increase. However, ozone-induced changes in plant cells at the gene level are almost unknown. The limited data available suggest close similarities between ozone-induced and pathogen-induced defence responses in plants. Several general defence genes that have been cloned in other studies will soon be applied to studies of gene expression in ozone-exposed plants. The use of molecular biological tools in ozone research should enable the development of highly specific and sensitive molecular markers for biomonitoring ozone-induced injuries in plants. 相似文献
148.
149.
Mutational and crystallographic analyses of the active site residues of the Bacillus circulans xylanase. 总被引:8,自引:7,他引:1
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W. W. Wakarchuk R. L. Campbell W. L. Sung J. Davoodi M. Yaguchi 《Protein science : a publication of the Protein Society》1994,3(3):467-475
Using site-directed mutagenesis we have investigated the catalytic residues in a xylanase from Bacillus circulans. Analysis of the mutants E78D and E172D indicated that mutations in these conserved residues do not grossly alter the structure of the enzyme and that these residues participate in the catalytic mechanism. We have now determined the crystal structure of an enzyme-substrate complex to 108 A resolution using a catalytically incompetent mutant (E172C). In addition to the catalytic residues, Glu 78 and Glu 172, we have identified 2 tyrosine residues, Tyr 69 and Tyr 80, which likely function in substrate binding, and an arginine residue, Arg 112, which plays an important role in the active site of this enzyme. On the basis of our work we would propose that Glu 78 is the nucleophile and that Glu 172 is the acid-base catalyst in the reaction. 相似文献
150.
Regeneration of catalytic activity of glutamine synthetase mutants by chemical activation: exploration of the role of arginines 339 and 359 in activity. 总被引:1,自引:0,他引:1
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A. M. Dhalla B. Li M. F. Alibhai K. J. Yost J. M. Hemmingsen W. M. Atkins J. Schineller J. J. Villafranca 《Protein science : a publication of the Protein Society》1994,3(3):476-481
In order to understand the nature of ATP and L-glutamate binding to glutamine synthetase, and the involvement of Arg 339 and Arg 359 in catalysis, these amino acids were changed to cysteine via site-directed mutagenesis. Individual mutations (Arg-->Cys) at positions 339 and 359 led to a sharp drop in catalytic activity. Additionally, the Km values for the substrates ATP and glutamate were elevated substantially above the values for wild-type (WT) enzyme. Each cysteine was in turn chemically modified to an arginine "analog" to attempt to "rescue" catalytic activity by covalent modification; 2-chloroacetamidine (CA) (producing a thioether) and 2,2'-dithiobis (acetamidine)(DTBA) (producing a disulfide) were the reagents used to effect these chemical transformations. Upon reaction with CA, both R339C and R359C mutants showed a significant regain of catalytic activity (50% and 70% of WT, respectively) and a drop in Km value for ATP close to that for WT enzyme. With DTBA, chemically modified R339C had a greater kcat than WT glutamine synthetase, but chemically modified R359C only regained a small amount of activity. Modification with DTBA was quantitative for each mutant and each modified enzyme had similar Km values for both ATP and glutamate. The high catalytic activity of DTBA-modified R339C could be reversed to that of unmodified R339C by treatment with dithiothreitol, as expected for a modified enzyme containing a disulfide bond. Modification of each cysteine-containing mutant to a lysine "analog" was accomplished using 3-bromopropylamine (BPA).(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献