Enzymes, reactions with

The final method of coupling enzyme reactions to electrochemistry is to immobilize an enzyme directly at the electrode surface. Enzyme electrodes provide the advantages already discussed for immobilization of enzymes. In addition, the transport of enzyme product from the enzyme active site to the electrode surface is greatly enhanced when the enzyme is very near to the electrode. The concept of combining an enzyme reaction with an amperometric probe should offer all of the advantages discussed earlier for ion-selective (potentiometric) electrodes with a much higher sensitivity. In addition, since the response of amperometric electrodes is linear, background can be selected. 

Zhang Y, Liu H, Yang W (2000) Free energy calculation on enzyme reactions with an efficient iterative procedure to determine minimum energy paths on a combined ab initio QM/MM potential energy surface. J Chem Phys 112 3483-3492... 

Figure 20.14 Periodate oxidation of HRP creates aldehyde groups on the carbohydrate chains of the enzyme. Reaction with a Fab fragment then may be done using reductive amination to produce a lower-molecular-weight complex than would be obtained using intact IgG antibodies.

The next stage leads to the transformation of the phosphorylated glyceric acid into pyruvic add, which is then converted into acetaldehyde and C02 by the enzyme carboxylase. As soon as the acetaldehyde is formed it enters into enzymic reaction with the phosphorylated glycer-aldehyde which is dehydrogenated to the acid while the aldehyde becomes ethyl alcohol. 

In the ground-breaking scientific paper that presented their work, Menten and Michaelis also derived an important mathematical formula. This formula describes the rate at which enzymes break down their substrates. It correlates the speed of the enzyme reaction with the concentrations of the enzyme and the substrate. Called the Michaelis-Menten equation, it remains fundamental to our understanding of how enzymes catalyze reactions. 

Nachtomi E. 1970. Metabolism of ethylene dibromide in the rat The enzymic reaction with glutathione iTLvitro and iTLvivQ. Biochem Pharmacol 19 2853-2860. 

To expand the cychc enzyme system and control the enzymic reaction with the use of an external inhibitor, and to examine the characteristics of this new system as an information-processing unit. 

Creatine phosphokinase activity has been reported to be minimally inhibited by hemolysis. Hemoglobin concentrations of 1.25 g/100 ml inhibit 5% and 2.5 g/100 ml, 12% (N5). However, in methods utilizing adenosine diphosphate in the reaction mixture, hemolysates containing 100 mg of hemoglobin per 100 ml may have apparent activities of 5-100 units/liter. The activity is presumably related to adenylate kinase in the erythrocyte (S33). In methods utilizing adenosine diphosphate in a coupled enzyme reaction with hexokinase and glucose-6-phosphatase, the inhibitory effect can be eliminated by adding sufficient adenosine mono-... 

The overall rate equation of complex single-route reaction with the linear detailed mechanism was derived and analyzed in detail by many researchers. King and Altman (1956) derived the overall reaction rate equation for single-route enzyme reaction with an arbitrary number of intermediates... 

Enzymic reactions with D-galactose oxidase or L-fucose isomerase were used for identification of the monosaccharide isolated after degradation of the esters. 

Immobilized enzyme beads of 0.6 cm diameter contain an enzyme that converts a substrate S to a product P by an irreversible unimolecular enzyme reaction with K = 0.012 kmol m and the maximum rate... 

The investigations of W. H. Stein and Moore and their colleagues were first reported in 1959 157). The inactivation of RNase by iodo-acetate was studied. A maximum in the rate of activity loss was noted at pH 5.5. Reaction with a methionine residue was found at pH 2.8 at pH 8.5-10 lysine residues were modified, but at pH 5.5-6.0 only histidine appeared to be involved. The specific reaction required the structure of the native enzyme. Reaction with histidine was not observed under a variety of denaturing conditions 158). Iodoacetamide did not cause activity loss, or only very slow loss, or alkylate His 119 in the native enzyme at pH 5.5. The negative charge on the carboxyl group of the iodoacetate ion was apparently essential. 

Several techniques have been developed for the determination of purine and pyrimidine derivatives in food sample and in particular for hypoxanthine quantification biosensors (220-223) and electrochemical methods making use of immobilized enzyme electrode (224 -227), electrochemical enzymatic-based HA methods (228,229), enzyme reaction with fluorimetric detection (230), radioimmunoassay (231), colorimetric methods (232), capillary electrophoresis (233), and TLC (234). Many HPLC methods have also been developed and are reported in Table 4 (235-247) the most recent ones are described next. 

Y. Sekine and E.A.H. Hall, A lactulose sensor based on coupled enzyme reactions with a ring electrode fabricated from tetrathiafulvalen-tetra-cyanoquinodimetane, Biosens. Bioelectron., 13(9) (1998) 995-1005. 

Enzyme Reactions with Incomplete Mass Transfer Influence of Immobilization 1113... 

Figure 7.1 Enzyme reactions with the cyano group.

The kinetics of enzyme reactions was first established by Michaelis and Menten, following the earlier work of Henri [23]. The famous Michaelis-Menten equation for the kinetics of an enzyme reaction with a single substrate is often written [23]... 

M 16] [P 16] The same findings as for the chemical reaction (see Chemical reaction in electrode dot devices with sequential voltage) were made for the luciferin-luciferase enzyme reaction with adenosine triphosphate on the six-phase electrode array device [100, 101]. The reaction rapidly followed the mixing, as evident from the luciferin luminescence of the droplet after merging. 

A method of great value in biochemistry for the verification of peak identities is the enzymatic peak shift technique.5 The approach utilizes the specificity of enzyme reactions with a nucleotide or class of nucleotides. The technique is especially useful in the characterization of nucleotides of cell extracts, because not only is the identity of the reactant verified, but so is the identity of the product formed. With the enzyme peak shift method, one aliquot of the sample is analyzed while a second aliquot is incubated with an excess of an enzyme that catalyzes a specific reaction involving the compound of interest. After the enzyme is deactivated, the second aliquot is chromatographed. 

Compare the initial rates (the first 10 min.) of the enzyme reactions with and without inhibitor. How many times slower was the reaction with inhibitor than without it ... 

Are these phenomena unique, or are they typical of biological systems From a mathematical perspective, enzyme systems fall into a class of nonlinear organization, and a chain of enzyme reactions with negative feedback easily can demonstrate oscillatory behavior [520]. Glass has noted that in general, any nonlinear system with multiple negative feedback may demonstrate oscillations that lead to chaotic behavior [595]. 

A, Chemical oxidation or enzyme reaction with HRP/H2O2 B, electrochemical reaction C, enzyme reaction with HRP/H202. 

Periappuram, C., Steinhauer, L., Barton, D.L., and Zon, J., 2000, The plastidic phosphoglucomutase from Arabidopsis. A reversible enzyme reaction with an important role in metabolic control. Plant Physiol. 122 1193-1200. 

Kinetic behavior becomes complicated when there are two chemical species that can both complex with the enzyme molecules. One of the species might behave as an inhibitor of the enzyme reaction with... 

The catalytic aspect of enzymes has prompted the development of new types of enzyme inactivators, which have been called suicide inhibitors . These inhibitors are substrates which are modified by masking the reactive functions. If they are unmasked by the catalytic action of the target enzyme, reaction of the reactive group with an active site or cofactor is made possible and this leads to the inactivation of the enzyme. Since the chemically reactive group is only liberated at the active site of the target enzyme, reactions with foreign molecules cannot occur. A number of examples of suicide inhibitors (or K -at inhibitors) can be found in a paper by Rando... 

In this test, oxygen from air is used for the enzymatic oxidation of (3-D-glucose in the presence of immobilized GOD to gluconic acid and hydrogen peroxide. Hydrogen peroxide can be determined by a second enzyme reaction. With horse radish peroxidase as catalyst o-phenylendiamine is oxidized by H2O2 to 2,3-diaminophenazine, which can be photometrically determined at 490 nm, thus establishing a quantitative relationship between active GOD sites and the intensity of the absorption band. 

The earliest quantitative theory of enzyme kinetics dates back to 1913, when Michaelis and Menten [27] succeeded in explaining a key feature of enzyme reactions with a very simple model. As an introduction and to establish the relationship between trace-level and bulk-species catalysis, this classical work and its subsequent refinements will now be reviewed. 

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