Showing papers on "Substrate (chemistry) published in 1979"

Fundamentals of Enzyme Kinetics

Abstract: Basic Principles of Chemical Kinetics Introduction to Enzyme Kinetics "Alternative" Enzymes Practical Aspects of Kinetics Deriving Steady-state Rate Equations Reversible Inhibition and Activation Tight-binding and Irreversible Inhibitors Reactions of More than One Substrate Use of Isotopes for Studying Enzyme Mechanisms Effect of pH on Enzyme Activity Temperature Effects on Enzyme Activity Regulation of Enzyme Activity Multienzyme Systems Fast Reactions Estimation of Kinetic Constants Standards for Reporting Enzymology Data Solutions and Notes to Problems Index

Rapid ethanol fermentation of cellulose hydrolysate. II. Product and substrate inhibition and optimization of fermentor design

Abstract: High concentrations of both ethanol and sugar in the fermentation broth inhibit the growth of yeast cells and the rate of product formation. Inhibitory effects of ethanol on the yeast strain Saccharomyces cerevisiae NRRL‐Y‐132 were studied in batch and continuous chemostat cultures. Growth was limited by either glucose or ethanol. Feed medium was supplemented with different ethanol concentrations. Ethanol was found to inhibit growth and the activity of yeast to produce ethanol in a noncompetitive manner. A linear kinetic pattern for growth and product formation was observed according to μ = μm (1 – P/Pm) and v = vm (1 – P/Pm′), where μm is the maximum specific growth rate at P = 0 (hr−1); Pm is the maximum specific product formation rate at P = 0 (hr−1); Pm is the maximum ethanol concentration above which cells do not grow (g/liter); Pm′ is the maximum ethanol concentration above which cells do not produce ethanol (g/liter). Substrate inhibition studies were carried out using short‐time experimental techniques under aerobic and anaerobic condition. The degree of substrate inhibition was found to be higher than that has been reported for ethanol fermentation of pure sugar. The kinetic relationships thus obtained were used to compute growth, substrate utilization, and alcohol production patterns and have been discussed with reference to batch and continuous fermentation of enzymatically produced bagasse hydrolysate.

Purification and properties of a prolyl endopeptidase from rabbit brain.

Abstract: — An enzyme with the specificity of a prolyl endopeptidase was purified about 880-fold from rabbit brain. The enzyme hydrolyzes peptidylprolyl-peptide and peptidylprolyl-amino acid bonds. Several biologically active peptides such as angiotensin, bradykinin, neurotensin. substance P and thyrotropin releasing hormone are degraded by hydrolysis of the bond between the carboxyl group of proline and the adjacent amino acid or ammonia respectively. The enzyme is activated by dithiothreitol and inhibited by heavy metals and thiol blocking agents. The serine protease inhibitor phenylmethanesulfonylfluoride has no effect on activity; however, inhibition was obtained with diisopropylfluorophosphate. Prolyl endopeptidase has a molecular weight of about 66,000 and a pH optimum of about 8.3. A new chromogenic substrate, N-benzyloxycarbonylglycyl-L-prolylsulfamethoxazole, was used for determination of enzyme activity. The substrate is hydrolyzed to N-benzyloxycarbonylglycyl-L-proline and free sulfamethoxazole which can be conveniently determined by a colorimetric procedure.

Transfer and detection of nucleic acids

Abstract: Improvements in the transfer and detection of separated nucleic acids, both RNA and DNA, are provided. For analysis of large DNA, the molecular weight segregated fractions of DNA are depurinated and fragmented to provide fractions having less than about 2 kb as single strands. With both RNA and DNA, the nucleic acid fractions are transferred after resolution to a chemically treated substrate and covalently affixed to the substrate. The resulting nucleotides affixed to the substrate are hybridized with labeled nucleotide probes and a volume exclusion agent, particularly a water soluble ionic polymer.

Comparison of substrate affinities among several rumen bacteria: a possible determinant of rumen bacterial competition.

Abstract: Five rumen bacteria, Selenomonas ruminantium, Bacteroides ruminicola, Megasphaera elsdenii, Streptococcus bovis, and Butyrivibrio fibrisolvens were grown in continuous culture. Estimates of substrate affinities were derived from Lineweaver-Burk plots of dilution rate versus substrate concentration. Each bacterium was grown on at least four of the six substrates: glucose, maltose, sucrose, cellobiose, xylose, and lactate. Wide variations in substrate affinities were seen among the substrates utilized by a species and among species for the same substrate. These wide differences indicate that substrate affinity may be a significant determinant of bacterial competition in the rumen where soluble substrate concentrations are often low. Growth of these bacteria in continuous culture did not always follow typical Michaelis-Menten kinetics. Inflated theoretical maximum growth rates and non-linear Lineweaver-Burk plots were sometimes seen. Maintenance energy expenditures and limitation of growth rate by factors other than substrate concentration (i.e., protein synthesis) are discussed as possible determinants of these deviations.

Specificity of guinea pig liver transglutaminase for amine substrates.

Abstract: The amine specificity of guinea pig liver transglutaminase, a model enzyme for endo-gamma-glutamine:epsilon-lysin transferases, was explored with the aid of synthetic substrates of high apparent affinities. As exemplified by dansyl- (5-dimethylamino-1-naphthalenesulfonyl), (2,4-dinitrobenzenesulfonyl)-, and (2,4,6-triisopropylbenzenesulfonyl)-cadaverines--each of which showed affinities of approximately 4 x 10(7) M-1--the best amine substrates carried a large hydrophobic substituent attached to an alkylamine side chain of about 7.2 A in length. Altogether, our results point to the importance of a hydrophobic binding region in the enzyme from where the alkyl side chain reaches into a narrow crevice toward the active center and positions the primary amine of the substrate for attacking the carbonyl group of the acyl enzyme intermediate.

Rhamnose-induced propanediol oxidoreductase in Escherichia coli: purification, properties, and comparison with the fucose-induced enzyme.

Abstract: Escherichia coli are capable of growing anaerobically on L-rhamnose as a sole source of carbon and energy and without any exogenous hydrogen acceptor. When grown under such condition, synthesis of a nicotinamide adenine dinucleotide-linked L-lactaldehydepropanediol oxidoreductase is induced. The functioning of this enzyme results in the regeneration of nicotinamide adenine dinucleotide. The enzyme was purified to electrophoretic homogeneity. It has a molecular weight of 76,000, with two subunits that are indistinguishable by electrophoretic mobility. The enzyme reduces L-lactaldehyde to L-1,2-propanediol with reduced nicotinamide adenine dinucleotide as a cofactor. The Km were 0.035 mM L-lactaldehyde and 1.25 mM L-1,2-propanediol, at pH 7.0 and 9.5, respectively. The enzyme acts only on the L-isomers. Strong substrate inhibition was observed with L-1,2-propanediol (above 25 mM) in the dehydrogenase reaction. The enzyme has a pH optimum of 6.5 for the reduction of L-lactaldehyde and of 9.5 for the dehydrogenation of L-1,2-propanediol. The enzyme is, according to the parameters presented in this report, indistinguishable from the propanediol oxidoreductase induced by anaerobic growth on fucose.

Secretion of cellulase and beta-glucosidase by Trichoderma viride ITCC-1433 in submerged culture on different substrates.

Abstract: Trichoderma viride ITCC-1433 produces high yields of cellulase and especially β-glucosidase when grown in submerged culture on different carbon sources. Cellulase synthesis was strongly repressed in the presence of glucose and only a low constitutive activity of β-glucosidase and carboxymethylcellulase, but no Avicelase, could be demonstrated when culturing T. viride on glucose. With carboxymethylcellulose (CMC) as a substrate the secretion of enzyme as well as growth depended on the degree of substitution, but in general CMC cannot be regarded either as a powerful inducer or as a carbon source. With insoluble cellulose, maximum enzyme production and activities were obtained using an alkali-treated cellulose powder. On this substrate the excretion of soluble protein into the culture broth increased and the protein concentration corresponded to cellulolytic activities.

Reaction of Azide Radicals with Amino Acids and Proteins

Abstract: SummaryThe azide radical N·3 reacts selectively with amino acids, in neutral solution preferentially with tryptophan (k(N·3 + TrpH) = 4·1 × 109 dm3 mol−1s−1) and in alkaline solution also with cysteine and tyrosine (k(N·3 + CyS−) = 2·7 × 109 dm3 mol−1s−1 and k(N·3 + TyrO−) = 3·6 × 109 dm3 mol−1s−1). Oxidation of the enzyme yADH by N·3 involves primary attacks, mainly at tryptophan residues, and subsequent slow secondary reactions. N·3-induced inactivation of yADH is likely to occur upon oxidation of tryptophan residues in the substrate binding pocket (58-TrpH and 93-TrpH) since the substrate ethanol, although unreactive with N·3, protects yADH and since elADH, which does not contain tryptophan in the substrate pocket, is comparatively resistant against N·3-attack. N·3 exhibits low reactivity with nucleic acid derivatives and it is inert towards aliphatic compounds such as methanol and sodium dodecylsulphate.