Molybdenum reductases

Compared with other transition metals in biological redox systems, the oxidation states likely to be used by molybdenum are very high (74). As discussed previously, the IV, V, and VI states are a likely set of participants in molybdenum oxidases, and while the II and III states remain viable for molybdenum reductases, it nevertheless seems likely that higher oxidation states will be found in these enzymes as well. Indeed, the substitution of tungsten for molybdenum in both nitrate reductase and nitrogenase indicates this likelihood as it is much more difficult to obtain the lower oxidation states of tungsten. 

The coupled proton-electron transfer mechanism can also be applied to the molybdenum reductases. For nitrate reductase, a scheme such as Reaction 20 is possible. A Mo (IV)-Mo (VI) couple is used to illustrate this, and while such a couple is viable for some nitrate reductases, the Mo(II)-Mo(IV) or the Mo(III)-Mo(V) couple could also be accommodated... 

The reductive biotransformation of drugs has been one of the least studied reactions, and many of the enzymes that are involved have not been well characterized. Some of the enzymes that catalyze reductive reactions of drugs are the cytochrome P450s, molybdenum reductases, alcohol dehydrogenases, carbonyl reductases, NADPH cytochrome P450 reductase, NAD(P)H— quinone oxidoreductases, and enzymes of the intestinal microflora (Matsunaga et al., 2006 Rosemond and Walsh, 2004). 

Molybdenum. Molybdenum is a component of the metaHoen2ymes xanthine oxidase, aldehyde oxidase, and sulfite oxidase in mammals (130). Two other molybdenum metaHoen2ymes present in nitrifying bacteria have been characteri2ed nitrogenase and nitrate reductase (131). The molybdenum in the oxidases, is involved in redox reactions. The heme iron in sulfite oxidase also is involved in electron transfer (132). 

The element molybdenum (atomic weight 95.95) constitutes 0.08% of the weight of nitrate reductase. If the molecular weight of nitrate reductase is 240,000, what is its likely quaternary structure ... 

Hydroxylamine, IV-benzoyl-lV-phenyl-in gravimetry, 1, 532 liquid-liquid extraction, 1, 544 Hydroxylamine, A -cinnamoyl-A -phenyl-liquid-liquid extraction, 1,544 Hydroxylamine, Ar,A -di-(-butyl-metal complexes, 2, 798 Hydroxylamine, Ay/V-diethyl-metal complexes, 2,798 Hydroxylamine, AAmethyl-metal complexes, 2,798 Hydroxylamine, A -2-naphthol-A -nitroso-ammonium salt — see Ncocupferron Hydroxylamine, A -nilrosophenyl-ammonium salt — see Cupferron Hydroxylamine ligands, 2, 101 Hydroxylamine oxido reductase, 6, 727 Hydroxylases molybdenum, 6,658,662 Hydroxylation arenes... 

The enzymes that utilize molybdenum can be grouped into two broad categories (1) the nitrogenases, where Mo is part of a multinu-clear metal center, or (2) the mononuclear molybdenum enzymes, such as xanthine oxidase (XO), dimethyl sulfoxide (DMSO) reductase, formate dehydrogenase (FDH), and sulfite oxidase (SO). The last... 

Sulfate reducers can use a wide range of terminal electron acceptors, and sulfate can be replaced by nitrate as a respiratory substrate. Molybdenum-containing enzymes have been discovered in SRB (also see later discussion) and, in particular, D. desulfuricans, grown in the presence of nitrate, generates a complex enzymatic system containing the following molybdenum enzymes (a) aldehyde oxidoreduc-tase (AOR), which reduces adehydes to carboxylic acids (b) formate dehydrogenase (FDH), which oxidizes formate to CO2 and (c) nitrate reductase (the first isolated from a SRB), which completes the enzy-... 

The molyhdopterin cofactor, as found in different enzymes, may be present either as the nucleoside monophosphate or in the dinucleotide form. In some cases the molybdenum atom binds one single cofactor molecule, while in others, two pterin cofactors coordinate the metal. Molyhdopterin cytosine dinucleotide (MCD) is found in AORs from sulfate reducers, and molyhdopterin adenine dinucleotide and molyb-dopterin hypoxanthine dinucleotide were reported for other enzymes (205). The first structural evidence for binding of the dithiolene group of the pterin tricyclic system to molybdenum was shown for the AOR from Pyrococcus furiosus and D. gigas (199). In the latter, one molyb-dopterin cytosine dinucleotide (MCD) is used for molybdenum ligation. Two molecules of MGD are present in the formate dehydrogenase and nitrate reductase. 

The molybdenum cofactor was liberated from D. gigas AOR, and under appropriate conditions was transferred quantitatively to nitrate reductase in extracts of Neurospora crassa nit-1 mutant) to yield active nitrate reductase 217). On the basis of molybdenum content, the activity observed for reconstitution with molybdenum cofactor of D. gigas was lower (25%) than the values observed for the procedure using extractable molybdenum cofactor of XO, used as reference. This result can now be put in the context of the difference in pterin present (MPT-XO and MCD-AOR) 218). 

D. desulfuricans is able to grow on nitrate, inducing two enzymes that responsible for the steps of conversion of nitrate to nitrite (nitrate reductase-NAP), which is an iron-sulfur Mo-containing enzyme, and that for conversion of nitrite to ammonia (nitrite reduc-tase-NIR), which is a heme-containing enzyme. Nitrate reductase from D. desulfuricans is the only characterized enzyme isolated from a sulfate reducer that has this function. The enzyme is a monomer of 74 kDa and contains two MGD bound to a molybdenum and one [4Fe-4S] center (228, 229) in a single polypeptide chain of 753 amino acids. FXAFS data on the native nitrate reductase show that besides the two pterins coordinated to the molybdenum, there is a cysteine and a nonsulfur ligand, probably a Mo-OH (G. N. George, personal communication). 

FPR studies at low temperature detect the presence of one iron-sulfur center and molybdenum. At low temperature a sample of nitrate reductase reduced by dithionite shows a rhombic signal (gm,x = 2.04, gmed = 1.94, and gnm = 1.90). This signal accounts for 0.84 spins/... 

Chlorate reductase has been characterized in strain GR-1 where it was found in the periplasm. It is oxygen-sensitive and contains molybdenum and [3Fe-4S] and [4Fe-4S] clusters (Kengen et al. 1999). 

McEwan AG, IP Ridge, CA McDevitt, P Hugenholtz (2002) The DMSO reductase family of microbial molybdenum enzymes molecular properties and the role in the dissimilatory reduction of toxic elements. 

Breese K, G Fuchs (1998) 4-hydroxybenzoyl-CoA reductase (dehydroxylating) from the denitrifying bacterium Thauera aromatica prosthetic groups, electron donor, and genes of a member of the molybdenum-flavin-iron-sulfur proteins. Eur J Biochem 251 916-923. 

Gibson J, M Dispensa, CS Harwood (1997) 4-hydroxybenzoyl coenzyme A reductase dehydroxylating is required for anaerobic degradation of 4-hydrozybenzoate by Rhodopseudomonas palustris and shares features with molybdenum-containing hydroxylases. J Bacterial 179 634-642. 

The chlorate reductase has been characterized in strain GR-1 where it was found in the periplasm, is oxygen-sensitive, and contains molybdenum, and both [3Fe-4S] and [4Fe-4S] clusters (Kengen et al. 1999). The arsenate reductase from Chrysiogenes arsenatis contains Mo, Fe, and acid-labile S (Krafft and Macy 1998), and the reductase from Thauera selenatis that is specific for selenate, is located in the periplasmic space, and contains Mo, Fe, acid-labile S, and cytochrome b (Schroeder et al. 1997). In contrast, the membrane-bound selenate reductase from Enterobacter cloacae SLDla-1 that cannot function as an electron acceptor under anaerobic conditions contains Mo and Fe and is distinct from nitrate reductase (Ridley et al. 2006). 

For nitrate reductase, evidence on the role of molybdenum in the catalytic mechanism of the enzyme from Neurospora was first presented in 1954 by Nicholas and Nason (21) and the position seems to have changed relatively little since then. The original conclusion (23) was that molybdenum functions as an electron carrier in the sequence ... 

So little is known about molybdenum enzymes other than milk xanthine oxidase that there is little to be said by way of general conclusions. In all cases where there is direct evidence (except possibly for xanthine dehydrogenase from Micrococcus lactilyticus) it seems that molybdenum in the enzymes does have a redox function in catalysis. For the xanthine oxidases and dehydrogenases and for aldehyde oxidase, the metal is concerned in interaction of the enzymes with reducing substrates. However, for nitrate reductase it is apparently in interaction with the oxidizing substrate that the metal is involved. In nitrogenase the role of molybdenum is still quite uncertain. 

Fig. 6.9 The catalysts for denitrification. Nitrate is reduced by a molybdenum enzyme while nitrite and oxides of nitrogen are reduced today mainly by copper enzymes. However, there are alternatives, probably earlier iron enzymes. The electron transfer bct complex is common to that in oxidative phosphorylation and similar to the bf complex of photosynthesis, while cytochrome c2 is to be compared with cytochrome c of oxidative phosphorylation. These four processes are linked in energy capture via proton (H+) gradients see Figure 6.8(a) and (b) and the lower parts of Fig. 6.9 which show separately the active site of the all iron NO-reductase, and the active site of cytochrome oxidase (02 reductase).

Figure 17.2 The structure of the pterin cofactor (1) which is common to most molybdenum- and tungsten-containing enzymes and schematic active site structures for members of the xanthine oxidase (2,3), sulfite oxidase (4) and DMSO reductase (5-7) enzyme families. (From Enemark et al., 2004. Copyright (2004) American Chemical Society.)...

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