Pyridine nucleotide oxidoreductase

Clostridium thermoaceticum contains the so-called AMAPOR (artificial-media-tor-accepting pyridine-nucleotide oxidoreductases), which are useful for electro-microbial regeneration of all four forms of pyridine nucleotides, too. An NADP(H) dependent AMAPOR from C. thermoaceticum has been purified and characterized [104]. It is able to react with rather different artificial mediators such as viologens or quinones, for example 1,4-benzoquinone, anthraquinone-2,6-disulfonate, or 2,6-dichloro-indophenol. 

Lactate dehydrogenase is a pyridine nucleotide oxidoreductase, a tetramer of 140 kD molecular weight, which has been extensively investigated (Bloxham et al., 1975 Eventoff et al., 1977). It catalyses the reversible oxidation of L-lactate to pyruvate using NAD+ as a coenzyme. The reaction scheme with a view of the active site with bound substrate and essential amino-acid side chains are depicted in Equation (3) and in Figure 17. The probable reaction mechanism, involving proton and hydride transfers,... 

IfE3 is a pyridine nucleotide independent reductase accepting electrons from artificial mediators, only this enzyme is necessary, if the mediators are reduced electrochemically. If they are reduced enzymicly El has to be included. Reduced pyridine nucleotides are formed by E2. These artificial mediator accepting pyridine nucleotide oxidoreductases (AMAPOR) catalyse Reaction [8] and [8a] forming NAD(P)H. Further examples are given in the text. Artificial mediators are indicated by med . Y are natural mediators except pyridine nucleotides. They may assist the electron transfer, but they are not necessary. 

USE OF ARTIFICIAL MEDIATOR ACCEPTING PYRIDINE NUCLEOTIDE OXIDOREDUCTASES (AMAPORs) FOR THE REGENERATION OF PYRIDINE NUCLEOTIDES... 

NADPH was regenerated under anaerobic conditions by crude extracts of C. thermoaceticum containing formate dehydrogenase (FDH) as well as artificial mediator accepting pyridine nucleotide oxidoreductases (AMAPORs) and formate as electron donor. For electromicrobial redox reactions see Section 6. 

Xanthobacter sp. strain Py2 may be grown with propene or propene oxide. On the basis of amino acid sequences, the monooxygenase that produces the epoxide was related to those that catalyzes the monooxygenation of benzene and toluene (Zhou et al. 1999). The metabolism of the epoxide is initiated by nucleophilic reaction with coenzyme M followed by dehydrogenation (Eigure 7.13a). There are alternative reactions, both of which are dependent on a pyridine nucleotide-disulfide oxidoreductase (Swaving et al. 1996 Nocek et al. 2002) ... 

Mammalian thioredoxin reductases are a family of selenium-containing pyridine nucleotide-disulfide oxidoreductases. These enzymes catalyze NADPH-dependent reduction of the redox protein thioredoxin (Trx), which contains a redox-active disulfide and dithiol group and by itself may function as an efficient cytosolic antioxidant [77]. One of the functions of Trx/ thioredoxin reductase system is the NADPH-catalyzed reduction of protein disulfide [78] ... 

The aldehyde dehydrogenases are members of a superfamily of pyridine nucleotide [NAD(P)+]-dependant oxidoreductases that catalyze the oxidation of aldehydes to... 

The pharmaceutical and fine chemical industry might use pure hydrogenase or partially purified enzyme preparations in bioconversion applications such as regio and stereoselective hydrogenation of target compounds (van Berkel-Arts et al. 1986). Enzymes are able to catalyse such stereospecific syntheses with ease. However, the cofactors for the NAD-dependent oxidoreductases are expensive. The pyridine nucleotide-dependent hydrogenases such as those from Ralstonia eutropha and hyperthermophilic archaea (Rakhely et al. 1999) make it possible to exploit H2 as a low-cost reductant. The use of inverted micelles in hydrophobic solvents, in which H2 is soluble, has advantages in that the enzymes appear to be stabilized. 

Pyridine nucleotide-dependent flavoenzyme catalyzed reactions are known for the external monooxygenase and the disulfide oxidoreductases However, no evidence for the direct participation of the flavin semiquinone as an intermediate in catalysis has been found in these systems. In contrast, flavin semiquinones are necessary intermediates in those pyridine nucleotide-dependent enzymes in which electron transfer from the flavin involves an obligate 1-electron acceptor such as a heme or an iron-sulfur center. Examples of such enzymes include NADPH-cytochrome P4S0 reductase, NADH-cytochrome bs reductase, ferredoxin — NADP reductase, adrenodoxin reductase as well as more complex enzymes such as the mitochondrial NADH dehydrogenase and xanthine dehydrogenase. 

As shown in the biosynthesis of granaticin, a hydride shift occurs intramolecularly. This process is mediated by an enzyme-bond pyridine nucleotide. A concerted abstraction of H-4 as a hydride in la and a C-5 deprotonation in 2a leads to the 4,5-enol ether 3a. The reduced form of the pyridine nucleotide transfers the hydride to C-6, simultaneously releasing a hydroxide to give 4a. Final tautomerization yields the dTDP-4-keto-6-deoxy-sugar in v-xylo configuration 4a. In other enzymes of the oxidoreductase type, the active site may show a different configuration. Thus, the intermediate 3a can be protonated from above at C-5 to yield the l-arabino isomer of 4a [2]. The stereochemistry of this mechanism was demonstrated by double labelling (cf. l-4b series), and as a net result proved a suprafacial 4—>6 hydride shift. 

Figure 12.18. Output of Pfam search results. Pfam search is performed with amino acid sequence derived from lipoamide dehydrogenase (Schizosaccharomyces pombe). The table for the trusted matches from Pfam-A for pyr redox (pyridine nucleotide disulfide oxidoreductase) and pyr redox dim (pyridine nucleotide disulfide oxidoreductase, dimerization) domains and their alignments (partial) to HMMs ( ->) are shown. The trusted matches from Pfam-B, the potential matches (Thi4 for thiamine biosynthetic enzyme domain), and the bead-on-a-string sketches are not shown. Select the linked domain name to view the functional description of the domain. The HMM alignments are followed by an option button (Align to seed or Align to family) that enables the user to view/save the multiple alignment of each matched family.

TrxRs are homodimeric flavoproteins [80] that catalyze the NADPH-dependent reduction of thioredoxin (Trx), a ubiquitous 12 kDa protein that is the major protein disulfide reductase in cells [81], and belongs to the pyridine nucleotide-disulfide oxidoreductase family [82]. Each monomer includes an FAD prosthetic group, a NADPH binding site and an active site containing a redox-active selenol group. Electrons are transferred from NADPH via FAD to the active-site selenol of TrxR, which then reduces the substrate Trx [83]. The crystal structure of TrxR is shown in Fig. 13 [84],... 

As is indicated in Table 5-3, P680, P70o> the cytochromes, plastocyanin, and ferredoxin accept or donate only one electron per molecule. These electrons interact with NADP+ and the plastoquinones, both of which transfer two electrons at a time. The two electrons that reduce plastoquinone come sequentially from the same Photosystem II these two electrons can reduce the two >-hemes in the Cyt b(f complex, or a >-heme and the Rieske Fe-S protein, before sequentially going to the /-heme. The enzyme ferre-doxin-NADP+ oxidoreductase matches the one-electron chemistry of ferredoxin to the two-electron chemistry of NADP. Both the pyridine nucleotides and the plastoquinones are considerably more numerous than are other molecules involved with photosynthetic electron flow (Table 5-3), which has important implications for the electron transfer reactions. Moreover, NADP+ is soluble in aqueous solutions and so can diffuse to the ferredoxin-NADP+ oxidoreductase, where two electrons are transferred to it to yield NADPH (besides NADP+ and NADPH, ferredoxin and plastocyanin are also soluble in aqueous solutions). 

The pyridine nucleotide-disulfide oxidoreductases, lipoamide dehydrogenase (4), glutathione reductase (5), and thioredoxin reductase (6-8) share so many properties in common that they will be compared and contrasted before being considered separately. As their group name implies, they catalyze the transfer of electrons between pyridine nucleotides and disulfides. In spite of their similarities they function in widely divergent metabolic roles. 

The gross structure of the pyridine nucleotide-disulfide oxidoreductases is the same, i.e., two polypeptide chains each containing a redox active... 

Amino Acid Analysis of Pyridine Nucleotide-Disulfide Oxidoreductases"... 

The specificity of glutathione reductase toward its disulfide substrate was emphasized in Section II,A, since there is virtually no reactivity with the substrates of the other pyridine nucleotide-disulfide oxidoreductases. Other authors have emphasized the lack of specificity of this enzyme since it can catalyze the reduction of a variety of mixed disulfides provided that glutathione or y-glutamylcysteine comprises one-half (IP5, 21%) Table IV summarizes these (39, 226-231). It is important to distin-... 

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