Cofactor NADH

Surface-modified electrodes were used for prevention of high overpotentials with direct oxidation or reduction of the cofactor, electrode fouling, and dimerization of the cofactor [7cj. Membrane electrochemical reactors were designed. The regeneration of the cofactor NADH was ensured electrochemically, using a rhodium complex as electrochemical mediator. A semipermeable membrane (dialysis or ultrafiltration) was integrated in the filter-press electrochemical reactor to confine... 

Ketoreductases catalyze the reversible reduction of ketones and oxidation of alcohols using cofactor NADH/NADPH as the reductant or NAD + /NADP+ as oxidant. Alcohol oxidases catalyze the oxidation of alcohols with dioxygen as the oxidant. Both categories of enzymes belong to the oxidoreductase family. In this chapter, the recent advances in the synthetic application of these two categories of enzymes are described. 

Kragl and Wandrey made a comparison for the asymmetric reduction of acetophenone between oxazaborolidine and alcohol dehydrogenase.[59] The oxazaborolidine catalyst was bound to a soluble polystyrene [58] and used borane as the hydrogen donor. The carbonyl reductase was combined with formate dehydrogenase to recycle the cofactor NADH which acts as the hydrogen donor. Both systems were run for a number of residence times in a continuously operated membrane reactor and were directly comparable. With the chemical system, a space-time yield of 1400 g L"1 d"1 and an ee of 94% were reached whereas for the enzymatic system the space-time yield was 88 g L 1 d"1 with an ee of >99%. The catalyst half-life times were... 

Hydrogen will not reduce ketones or imines using CATHy or related catalysts. Inorganic hydrogen donors that have been used include dithionite and di-hydrogenphosphite salts, metal hydrides such as sodium borohydride, and sodium cyanoborohydride. Recently, amines have been shown to function as hydrogen donors with some catalysts. The enzymic cofactor NADH can be used stoichiometrically, and the potential exists to use this catalytically [56]. 

An immobilized-enzyme continuous-flow reactor incorporating a continuous direct electrochemical regeneration of NAD + has been proposed. To retain the low molecular weight cofactor NADH/NAD+ within the reaction system, special hollow fibers (Dow ultrafilter UFb/HFU-1) with a molecular weight cut-off of 200 has been used [32],... 

Since many of the transformations undergone by metabolites involve changes in oxidation state, it is understandable that cofactors have been developed to act as electron acceptors/ donors. One of the most important is that based on NAD/NADP. NAD+ can accept what is essentially two electrons and a proton (a hydride ion) from a substrate such as ethanol in a reaction catalysed by alcohol dehydrogenase, to give the oxidized product, acetaldehyde and the reduced cofactor NADH plus a proton (Figure 5.2). Whereas redox reactions on metal centres usually involve only electron transfers, many oxidation/reduction reactions in intermediary metabolism, as in the case above, involve not only electron transfer but... 

The three-dimensional X-ray structure of the enzyme [19] reveals that several Thr residues occur in both the NADH cofactor and substrate binding sites (Fig. 21.5 A see p. 463). A Met residue (Metl7) is also present at the interface between the cofactor NADH and a substrate analog pyridine-2,6-dicarboxylate (PDC) (Fig. 21.5 A). Therefore, we prepared a sample of DHPR that was selectively labeled in these amino acid residues as follows 13C /1H Met, 13C /1H lie, 13C/1H Thr and uniformly 2H-labeled elsewhere ([MIT]-DHPR). This labeling can be achieved by supplementing the media with appropriate commercially available labeled amino acids, 12C/2H-labeled glucose and DzO [20] (see also the caption to Fig. 21.5 for details). 

Just as in the preceding examples, early indications of tunneling in enzyme-catalyzed reactions depended on the failure of experiments to conform to the traditional expectations for kinetic isotope effects (Chart 3). Table 1 describes experimental determinations of -secondary isotope effects for redox reactions of the cofactors NADH and NAD. The two hydrogenic positions at C4 of NADH are stereochemically distinct and can be labeled individually by synthetic use of enzyme-catalyzed reactions. In reactions where the deuterium label is not transferred (see below), an... 

Determination of the Michaelis constant for the cofactor NADH (A m.NADn) was carried out by measuring the initial rate of oxidation of NADH as a function of its concentration, at a constant concentration of pyruvate. All the solutions were prepared in 0.1 M phosphate buffer pH 7.55. 

In the second approach the reducing equivalents are suppHed by a nicotinamide cofactor (NADH or NADPH) and for commercial viability it is necessary to regenerate the cofactor using a sacrificial reductant ]12]. This can be achieved in two ways substrate coupled or enzyme coupled (Scheme 6.2). Substrate-coupled regeneration involves the use of a second alcohol (e.g. isopropanol) that can be accommodated by the KRED in the oxidative mode. A problem with this approach is that it affords an equilibrium mixture of the two alcohols and two ketones. In order to obtain a high yield of the desired alcohol product a large excess of the sacrificial alcohol needs to be added and/or the ketone product (acetone) removed... 

A Zn2+ at the active site polarizes the carbonyl oxygen of acetaldehyde, allowing transfer of a hydride ion (red) from the reduced cofactor NADH. The reduced intermediate acquires a proton from the medium (blue) to form ethanol. Alcohol Dehydrogenase Mechanism... 

Ketoreductases (KREDs) are dependent on nicotinamide cofactors NADH or NADPH. Due to the reaction mechanism, these rather costly cofactors are needed in stoichiometric amounts, disclosing an economic problem that has to be dealt with when using these enzymes. Many different possibilities for cofactor recycling have been established with three major approaches finding application in research and industry (Fig. 13). Further regeneration systems, such as electrochemical methods, are not discussed within this review [22-24, 37, 106-108],... 

Chemically modified electrodes (CMEs) for electrocatalytic oxidation of the reduced form of the nicotinamide adenine dinucleotide cofactor (NADH) are discussed. The work of the authors in the field is reviewed. CMEs based on adsorbed polyaromatic redox mediators (phenoxazines and phenothiazines) and the deposition of aqueous insoluble redox polymers are described. 

Scheme 4.101). The cofactor NADH was regenerated by the simultaneous oxidation of isopropanol to acetone. 

Answer Glycerol 3-phosphate and dihydroxyacetone 3-phosphate differ only at C-2. A dehydrogenase with the cofactor NADH acting on dihydroxyacetone 3-phosphate would form glycerol 3-phospate. 

Answer The measurement of the activity of alanine aminotransferase by measurement of the reaction of its product with lactate dehydrogenase is an example of a coupled assay. The product of the transamination (pyruvate) is rapidly consumed in the subsequent indicator reaction, catalyzed by an excess of lactate dehydrogenase. The dehydrogenase uses the cofactor NADH, the disappearance of which is conveniently measured by observing the rate of decrease in NADH absorption at 340 nm. Thus, the rate of disappearance of NADH is a measure of the rate of the aminotransferase reaction, if NADH and lactate dehydrogenase are added in excess. 

Because reduced redox cofactors, NADH and FADH2, are produced in the mitochondria, there is no need for shuttle mechanisms to reoxidize them via oxidative phosphorylation. NADH is reduced directly by complex I. FADH2 is reduced by the electron transfer flavoprotein, which then reduces ubiquinone. See Chapter 17 for details. 

Fig. 34 Enzymatic reduction in biphasic media. Ketones are reduced enantioselective to the corresponding (S)-alcohols by ADH from Rhodococcus erythropolis. Regeneration of the cofactor NADH is carried out by FDH from C. boidinii. The introduction of a biphasic system allows higher substrate concentrations...

There are many carrier molecules for electrons one is called the nicotinamide adenine dinucleotide (NAD+) and another is the flavin adenine dinucleotide FAD+. The reduced cofactors NADH and FADH2 transfer electrons to the electron transport chain. FMN receives electrons from NADH and passes them to coenzyme Q through Fe-S systems. Coenzyme Q receives electrons from FMN and FADH2 through Fe-S systems. Cytochromes receive electrons from the reduced form of coenzyme Q. Each cytochrome consists of a heme group, and the iron of the heme group is reduced when the cytochrome receives an electron Fe3+ Fe2+. At the end of the electron transfer chain, oxygen is reduced to water. 

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