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排序方式: 共有449条查询结果,搜索用时 15 毫秒
141.
Ferrocene-mediated carbon paste electrode modified with d-fructose dehydrogenase for batch mode measurement of d-fructose 总被引:1,自引:0,他引:1
A mediated modified carbon paste and renewable surface electrode for fructose amperometric measurement based on d-fructose dehydrogenase (FDH) was prepared and optimized. Commercially available ferrocene (FcH) and hydroxymethyl ferrocene
(FcCH2OH) were used as mediators. The substituted FcH showed better linearity and higher sensitivity. The influence of different
experimental parameters was studied for optimum analytical performance. The final FDH-modified electrode showed good analytical
performance for batch mode measurements of fructose. 相似文献
142.
The kinetics of free glucose-6-phosphate dehydrogenase (G-6-PDH), biotinylated G-6-PDH, and biotinylated G-6-PDH complexed
with avidin were investigated. The kinetics of the free enzyme were consistent with a sequential rather than a ping-pong mechanism.
The kinetics of the biotinylated enzyme were similar to that of the free enzyme, but the kinetic constants were different;
theK
m value for NADP was halved, whereas theK
m for G-6-P decreased only slightly. In the presence of avidin, theK
m of biotinylated G-6-PDH for G-6-P nearly doubled whereas theK
m for NADP did not change significantly. Avidin complexed with biotinylated G-6-PDH inhibited the enzyme from acting. Based
upon these reactions, it was possible to devise assays for either free biotin or free avidin using biotinylated G-6-PDH as
the indicator enzyme. Concentrations of biotin between 40 and 60 mg/mL, or of 25–95 Μg/mL of avidin could be measured within
2 min through the use of biotinylated G-6-PDH. 相似文献
143.
α-Amino acids and α-keto acids are versatile building blocks for the synthesis of several commercially valuable products in the food, agricultural, and pharmaceutical industries. In this study, a novel transamination-like reaction catalyzed by leucine dehydrogenase was successfully constructed for the efficient enzymatic co-synthesis of α-amino acids and α-keto acids. In this reaction mode, the α-keto acid substrate was reduced and the α-amino acid substrate was oxidized simultaneously by the enzyme, without the need for an additional coenzyme regeneration system. The thermodynamically unfavorable oxidation reaction was driven by the reduction reaction. The efficiency of the biocatalytic reaction was evaluated using 12 different substrate combinations, and a significant variation was observed in substrate conversion, which was subsequently explained by the differences in enzyme kinetics parameters. The reaction with the selected model substrates 2-oxobutanoic acid and L-leucine reached 90.3% conversion with a high total turnover number of 9.0 × 106 under the optimal reaction conditions. Furthermore, complete conversion was achieved by adjusting the ratio of addition of the two substrates. The constructed reaction mode can be applied to other amino acid dehydrogenases in future studies to synthesize a wider range of valuable products. 相似文献
144.
Hüsing Nicola Reisler Emil Zink Jeffrey I. 《Journal of Sol-Gel Science and Technology》1999,15(1):57-62
Glutamate dehydrogenase is encapsulated in a transparent porous silicate matrix by using sol-gel techniques. The inorganic polymer is formed around the enzyme (MW > 300,000 D). The enzyme is active in the material, catalyzes the reaction of L-glutamate to 2-oxoglutarate and follows Michaelis-Menten kinetics. The allosteric regulators ADP and GTP inhibit or activate the reaction; at pH 6, GTP acts as a strong activator and ADP acts as an inhibitor. This system involves a complex series of interactions; the co-enzyme NAD+ is required for catalysis, large-scale conformational changes accompany the binding of the substrate and coenzyme to the enzyme, the activators/inhibitors must bind to the enzyme to regulate the reactions, and the substrates and products must diffuse through the matrix to and from the binding site. The influence of the unique matrix on the complex enzymatic system is discussed. 相似文献
145.
N. A. Willoughby T. Kirschner M. P. Smith R. Hjorth N. J. Titchener-Hooker 《Journal of chromatography. A》1999,840(2):375-204
Alcohol dehydrogenase (ADH) from solutions of homogenised packed bakers’ yeast has been successfully purified using immobilised metal-ion affinity chromatography in an expanded bed. Method scouting carried out using pure ADH solutions loaded onto 5-ml HiTrap columns charged with Zn2+, Ni2+ and Cu2+ and eluted using 0–50 mM EDTA gradient found that charging with Zn2+ gave the highest recovery and the lowest EDTA concentration required for elution. These results were used to develop a protocol for the expanded bed system and further tested using clarified yeast homogenate loaded onto XK16/20 packed beds (approximately 30 ml) packed with Chelating Sepharose FastFlow matrix in order to determine the optimum elution conditions using EDTA. The ADH was found to elute at 5 mM EDTA and the dynamic and total binding capacities of Streamline chelating for ADH were found to be 235 U/ml and 1075 U/ml matrix, respectively. Expanded bed work based on a step EDTA elution protocol demonstrated that ADH could be successfully eluted from unclarified homogenised bakers’ yeast diluted to 10 mg/ml total protein content with a recovery of 80–100% that was maintained over five consecutive runs with a vigorous clean-in-place procedure between each run. 相似文献
146.
Alain Cartier David Brown Bernard Maigret Sandrine Boschi-Muller Sophie Rahuel-Clermont Guy Branlant 《Theoretical chemistry accounts》1999,101(1-3):241-245
In the framework of a theoretical approach to the relationship between structure and reactivity of the catalytic centers
of enzymes, glyceraldehyde-3 phosphate dehydrogenase (GAPDH) has been chosen as a model enzyme. In GAPDH, the proximity of
His176 increases the reactivity of Cys149 at neutral pH; however, its presence alone is not sufficient to explain the reactivity of the catalytic Cys. In order to
determine which other interactions play an important role, a study of the geometric and electronic structure of the catalytic
site has been made using a hybrid quantum mechanics/molecular mechanics local self-consistent field method. This allows the
computation of the electronic properties of amino acid residues in subsystems influenced by other parts of the macromolecule.
The quantum subsystem was centered on the Cys149 residue of GAPDH. The structures of GAPDH taken from the crystallographic database did not include hydrogen atoms and these
had to be added taking into account the fact that, in the active site, His176 has three tautomeric forms: δ-His protonated, ε-His protonated and His+. The results presented here suggest that the most stable His…Cys system in GAPDH is a strongly hydrogen-bonded Cys149
−/His176
+ ion pair.
Received: 24 March 1998 / Accepted: 3 September 1998 / Published online: 23 November 1998 相似文献
147.
148.
Alves Lourdes A. Vitolo Michele Felipe Maria das Graças A. de Almeida e Silva João Batista 《Applied biochemistry and biotechnology》2002,98(1-9):403-413
The sugarcane bagasse hydrolysate, which is rich in xylose, can be used as culture medium for Candida guilliermondii in xylitol production. However, the hydrolysate obtained from bagasse by acid hydrolysis at 120°C for 20 min has by-products
(acetic acid and furfural, among others), which are toxic to the yeast over certain concentrations. So, the hydrolysate must
be pretreated before using in fermentation. The pretreatment variables considered were: adsorption time (15,37.5, and 60 min),
type of acid used (H2So4 and H3Po4), hydrolysate concentration (original, twofold, and fourfold. concentrated), and active
charcoal (0.5, 1.75 and 3.0%). The suitability of the pretreatment was followed by measuring the xylose reductase (XR) and
xylitol dehydrogenase (XD) activity of yeast grown in each treated hydrolysate. The response surface methodology (24 full factorial design with a centered face) indicated that the hydrolysate might be concentrated fourfold and the pH adjusted
to 7.0 with CaO, followed by reduction to 5.5 with H3PO4. After that it was treated with active charcoal (3.0%) by 60 min. This pretreated hydrolysate attained the high XR/XD ratio
of 4.5. 相似文献
149.
《Analytical letters》2012,45(12):2429-2439
Abstract A combination of enzyme‐based spectrophotometric analysis and alkaline hydrolysis was developed for the measurement of poly(3‐hydroxybutyrate) (PHB). The principle of the determination is as follows: alkaline hydrolysis decomposes PHB into its monomer product 3‐hydroxybutyrate, which is followed with enzymatic reaction catalyzed by 3‐hydroxybutyrate dehydrogenase in the presence of nicotinamide adenosine dinucleotide (NAD). The product, nicotinamide adenosine dinucleotide with hydrogen (NADH) results in a spectrophotometric signal at 340 nm. This method shows high performance characteristics with simple operations. 相似文献
150.
Summary. The ability of the muscular carnitine pool to accept and temporally donate acetyl groups (from and towards the coenzyme A pool) is an important functional role of carnitine within biological systems that is often overlooked within the scientific literature. The present review will discuss recent research demonstrating the existence of a period of inadequate acetyl-CoA delivery towards the tricarboxylic acid cycle (the so-called ‘acetyl group deficit’), which occurs as a consequence of the impaired integration of cytosolic (glycolysis) and mitochondrial energy producing pathways at the onset of muscular contraction; due to a lag in the activation of the pyruvate dehydrogenase complex. During this period of inadequate acetyl-CoA delivery, acetyl groups can be sequestered from the limited muscular acetylcarnitine reserve in an attempt to sustain continued tricarboxylic acid cycle flux. Following on from this, the present review will highlight the metabolic and functional benefits to be gained by overcoming this period of metabolic inertia, through elevating the concentration of acetylcarnitine prior to physical exercise; in the presence and absence of pyruvate dehydrogenase complex activation and through appropriately timed ‘warm-up’ exercise. 相似文献