NADH/NADPH cofactors enzymes

Oxidoreductases these enzymes catalyze redox reactions. Examples are oxidases that catalyze oxidation of a substrate by reducing molecular oxygen (02), and peroxidases that reduce H202. Laccases (EC 1.10.3.2) are oxidases that catalyze the oxidation of (poly)phenolic substrates. Reductases and dehydrogenases (EC 1.1.1) catalyze the reduction of carbonyls, using NADH/NADPH cofactors. Catalases (EC 1.11.1.6) catalyze the decomposition of H202 to 02 and H20. 

Oxidoreductases are a family of enzymes that catalyze a number of industrially important reactions, but they often require additional nicotinamide (NADH or NADPH) cofactors which... 

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. 

Reduced flavins (FADH2, FMNH2, and riboflavin) generated by flavin-dependent reductases have been hypothesized to reduce azo dyes in a nonspecific chemical reaction, and flavin reductases have been revealed to be indeed anaerobic azoreductases. Other reduced enzyme cofactors, for example, NADH, NADH, NADPH, and an NADPH-generating system, have also been reported to reduce azo dyes. Except for enzyme cofactors, different artificial redox mediating compounds, especially such as quinines, are important redox mediators of azo dye anaerobic reduction (Table 1). 

Heterofermentative LAB have the capability to utilize high concentrations of fructose such that the mannitol concentration in the fermentation broth could reach more than 180g/L, which is enough to be separated from the cell-free fermentation broth by cooling crystallization. Lactic and acetic acids can be recovered by electrodialysis (Soetaert et al., 1995). The enzyme mannitol dehydrogenase responsible for catalyzing the conversion of fructose to mannitol requires NADPH (NADH) as cofactor. Thus, it is possible to develop a one-pot enzymatic process for production of mannitol from fructose if a cost-effective cofactor regeneration system can be developed (Saha, 2004). The heterofermentative LAB cells can be immobilized in a suitable support, and... 

But the application of redox enzymes is also connected with intrinsic problems. The difficulty with redox enzymes in synthetic processes is based on the fact that this class of biocatalysts is dependent on freely dissociated (like NADH, NADPH, pteridin) or enzyme-bound (like FMN, FAD, thio-tryosine, PQQ, or cytochrome) cofactors, respectively, prosthetic groups (Fig. 1) in stoichiometric amounts to shuttle the redox equivalents from the enzyme to the substrate. 

In the oxidation process of an alcohol by NAD+ (or NADP+), the enzymatic base positioned above the carbonyl takes back its proton, and the electrons in the O—H bond shift down and push out the hydride, which is immediately accepted by C-4 of NAD+ (or NADP+). The products are just what they were started out with, a ketone and NADH (or NADPH) (Figure 1.39). The mechanism of oxidation of amines to imines as well as of aldehyde to carboxylate is similar to the oxidation of alcohol. In the reduction process of a ketone by NADH (or NADPH), both the ketone substrate and cofactor are bound in the enzyme s active site, and C-4 of the nicotinamide ring is positioned very close to the carbonyl carbon of the ketone. As an enzymatic group transfers a proton to the ketone oxygen, the carbonyl carbon becomes more electrophilic and is attacked by a hydride from NADH (or NADPH). The ketone is reduced to an alcohol, and the NADH or NADPH cofactor is oxidized to NAD+ or... 

The first indication of the existence of so-called Baeyer-Villiger monooxygenases (BVMOs) was reported in the late 1940s [56]. It was observed that certain fungi were able to oxidize steroids via a BV reaction [56], but two decades elapsed before the first BVMOs were isolated and characterized [57, 58]. All characterized BVMOs contain a flavin cofactor that is vital for the catalytic activity of the enzyme, Furthermore, NADH or NADPH cofactors are needed as electron donors. Careful inspection of all available biochemical data on BVMOs has revealed that at least two discrete classes of BVMOs exist, types I and II [59]. 

Recombinant Saccharomyees cerevisiae, able to ferment the pentoses D-xylose and L-arabinose, was modified for improved fermentation rates and yields. Pentose fermentation is relevant when low cost raw materials such as plant hydrolysates are fermented to ethanol. The two most widespread pentose sugars in our biosphere are D-xylose and L-arabinose. S. cerevisiae is unable to ferment pentoses but has been engineered to do so however rates and yields are low. The imbalance of redox cofactors (excess NADP and NADH are produced) is considered a major limiting factor. For the L-arabinose fermentation we identified an NADH-dependent L-xylulose reductase replacing the previously known NADPH-dependent enzyme. For D-xylose fermentation we introduced an NADP-dependent glyceraldehyde 3-phospate dehydrogenase to regenerate NADPH. 

Most asymmetric reductions that can be enzymatically effected have been the reactions of ketones. These reactions can be conducted with whole cells as well as with isolated enzymes. In the latter case, of course, at least one equivalent of a cofactor such as NADH or NADPH (nicotinamide adenine dinucleotide) is required to serve as the actual reductant in the reaction system. 

Indicine IV-oxide (169) (Scheme 36) is a clinically important pyrrolizidine alkaloid being used in the treatment of neoplasms. The compound is an attractive drug candidate because it does not have the acute toxicity observed in other pyrrolizidine alkaloids. Indicine IV-oxide apparently demonstrates increased biological activity and toxicity after reduction to the tertiary amine. Duffel and Gillespie (90) demonstrated that horseradish peroxidase catalyzes the reduction of indicine IV-oxide to indicine in an anaerobic reaction requiring a reduced pyridine nucleotide (either NADH or NADPH) and a flavin coenzyme (FMN or FAD). Rat liver microsomes and the 100,000 x g supernatant fraction also catalyze the reduction of the IV-oxide, and cofactor requirements and inhibition characteristics with these enzyme systems are similar to those exhibited by horseradish peroxidase. Sodium azide inhibited the TV-oxide reduction reaction, while aminotriazole did not. With rat liver microsomes, IV-octylamine decreased... 

Unlike the whole-cell system, enzymatic reductions require the addition of a hydride donating cofactor to regenerate the reduced form of the enzyme. Depending on the chosen ADH, the cofactor is usually NADH or NADPH, both of which are prohibitively expensive for use in stoichiometric quantities at scale. Given the criticality of cofactor cost, numerous methods of in situ cofactor regeneration, both chemical and biocatalytic, have been investigated. However, only biocatalytic regeneration has so far proven to be sufficiently selective to provide the cofactor total turnover numbers of at least 10 required in production. 

This enzyme [EC 1.14.13.11], also known as cinnamate 4-hydroxylase, catalyzes the reaction of frans-cinnamate with NADPH and dioxygen to generate 4-hydroxycmna-mate, NADP+, and water. The enzyme, which uses a heme-thiolate as a cofactor, can also replace NADPH with NADH (however, the reaction will proceed slower). 

Nitrate reductase (NADH) [EC 1.6.6.1], also known as assimilatory nitrate reduetase, eatalyzes the reaction of NADH with nitrate to produee NAD+, nitrite, and water. This enzyme uses FAD or FMN, heme, and a molybdenum ion as eofaetors. (2) Nitrate reductase (NAD(P)H) [EC 1.6.6.2], also known as assimilatory nitrate reduetase, eatalyzes the reaetion of NAD(P)H with nitrate to produee NAD(P)+, nitrite, and water. This enzyme uses FAD or FMN, heme, and a molybdenum ion as eofaetors. (3) Nitrate reductase (NADPH) [EC 1.6.6.3] eatalyzes the reaetion of NADPH with nitrate to produee NADP+, nitrite, and water. This enzyme uses FAD, heme, and a molybdenum ion as cofactors. (4) Nitrate reduetase (eytoehrome) [EC 1.9.6.1] catalyzes the reaetion of nitrate with ferroeytochrome to produce nitrite and ferrieytoehrome. (5) Nitrate reductase (ac-eeptor) [EC 1.7.99.4], also known as respiratory nitrate... 

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... 

I I 3. The answer is c. (Hardman, pp 1243-1247.) Antimetabolites of folic acid such as methotrexate, which is an important cancer chemotherapeutic agent, exert their effect by inhibiting the catalytic activity of the enzyme dihydrofolate reductase. The enzyme functions to keep folic acid in a reduced state. The first step in the reaction is the reduction of folic acid to 7,8-dihydrofolic acid (FH2), which requires the cofactor nicotinamide adenine dinucleotide phosphate (NADPH). The second step is the conversion of FH2 to 5,6,7,8-tetrahydrofolic acid (FH ). This part of the reduction reaction requires nicotinamide adenine dinucleotide (NADH) or NADPH. The reduced forms of folic acid are involved in one-carbon transfer reactions that are required during the synthesis of purines and pyrimidine thymidylate. The affinity of methotrexate for dihydrofolate reductase is much greater than for the substrates of folic acid and FH2. The action of... 

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