Molybdenum enzymes reduction potentials

Although molybdenum and tungsten enzymes carry the name of a single substrate, they are often not as selective as this nomenclature suggests. Many of the enzymes process more than one substrate, both in vivo and in vitro. Several enzymes can function as both oxidases and reductases, for example, xanthine oxidases not only oxidize purines but can deoxygenate amine N-oxides [82]. There are also sets of enzymes that catalyze the same reaction but in opposite directions. These enzymes include aldehyde and formate oxidases/carboxylic acid reductase [31,75] and nitrate reductase/nitrite oxidase [83-87]. These complementary enzymes have considerable sequence homology, and the direction of the preferred catalytic reaction depends on the electrochemical reduction potentials of the redox partners that have evolved to couple the reactions to cellular redox systems and metabolic requirements. 

Table 3b Reduction Potentials for the Molybdenum Sites in Several Enzymes...

The postulated catalytic cycles for pterin-containing molybdenum enzymes involve a two-electron change at the molybdenum atom (Mo(VI) Mo(IV)). Microcoulometric titrations of nitrate reductase Chlorella vulgaris) (76), milk xanthine oxidase (77), and sulfite oxidase (78) show that their molybdenum centers are reduced by two electrons. The reduction potentials for the molybdenum center of chicken liver sulfite oxidase are strongly dependent upon pH and upon anion concentration (78). 

Xanthine is converted to uric acid at the molybdenum center of the enzyme, and the electrons are removed from the enzyme by oxidation of the flavin center. From early reductive titrations of xanthine oxidase with sodium dithionite, it was proposed that reducing equivalents were equilibrated among the four redox-active centers (Mo-co, two separate Fe2S2 centers, flavin) at a rate that was rapid relative to the overall catalytic rate of substrate turnover (243). Under such conditions, the flux of reducing equivalents through the enzyme should be influenced by the relative reduction potentials of the redox centers involved (244). Any effects of pH and temperature on the reduction potentials of individual redox components would affect the apparent rates of intramolecular transfer of the enzyme. 

Oxidation-Reduction Potentials Associated with Molybdenum in Enzymes... 

The iron-sulfur cubes that have been detected by x-ray analysis (49) as constituents of the enzymes ferridoxin and high-potential iron protein have been extruded from these enzymes by replacing the sulf-hydryl ligands of the enzymes with simple mercaptans, and these cubes identified with the corresponding synthetic compounds, II (50, 51, 52, 53). The latter have oxidation-reduction properties that closely mimic those of the enzymes. Similarly, an iron-sulfur-molybdenum double cube has been ejected from nitrogenase, and a similar double cube has been synthesized by Holm and his collaborators (54). It remains to be seen whether or not the iron-sulfur-molybdenum double cube can mimic the properties of nitrogenase (55). 

Nitrate reductases are found in a wide range of eukaryotes and prokaryotes and have a crucial role in nitrogen assimilation and dissimilation (see Chapter 8.14). These enzymes catalyze the reaction shown in Equation (5) for the assimilatory nitrate reductases, this is followed by the reduction of nitrite to ammonia. Dissimilatory nitrate reductases [142 147] catalyze the reduction of nitrate to nitrite for respiration, to generate a transmembrane potential gradient.The assimilatory nitrate reductases have a molybdenum center similar to that of sulfite oxidase (see... 

Nitrogenase, as must now become clear, is a complex enzyme of two component proteins which requires ATP, a reductant, a reducible substrate, Mg " " as an activator, and an anaerobic environment to function. To this complexity must be added the difficulty that the component proteins have no enzymatic half reactions . There are, perhaps, four main questions to decide about the mechanism (1) the role(s) of the two component proteins (2) the role(s) of ATP (3) the nature of the active site(s) and (4) the mechanism of N2 reduction. Despite the complexities and difficulties mentioned above, progress in the last 15 years has partly answered all these questions. The Fe protein mediates an ATP-dependent electron transfer from the donor to the MoFe protein which contains the active site. MgATP binds and induces a conformational change in the Fe protein which lowers its redox potential. FeMoco, the molybdenum cofactor, which may be part of the active site of N2 reduction, has been isolated and partly characterized, while an intermediate in N2 reduction has recently been discovered (Thorneley ct al., 1978). The next part of this chapter describes the evidence for these claims. This evidence involves the noncatalytic reactions of the individual proteins, their... 

Oxomolybdenum centers are present in a series of enzymes known as molybdenum hydroxylases [4] which catalyze oxygen atom transfer reactions of a variety of small molecules. Examples include the oxidation of xanthine to uric acid and the reduction of N03 to N02"- The reactions are equivalent to two-electron transfers, and the molybdenum atom cycles between oxidation states VI and IV during these processes. Molybdenum (VI) typically contains two multiply bonded oxo (or sulfido) groups, and molybdenum (IV) contains one. Therefore, it is believed that Mo02 and MoO +centers intercovert by coupled transfer of protons and electrons over a narrow potential interval during turnover of these enzymes. However, other explanations are possible. 

Table II summarizes all currently available data on the redox potentials of molybdenum in enzymes and other molybdenum-containing proteins. In all cases the values quoted were obtained by estimation, by EPR, of the fractional conversion of molybdenum to Mo(V) in a series of samples poised at known potentials. These potentials were measured with a platinum electrode, using a variety of low-molecular-weight dyes as mediators of the oxidation-reduction processes and usually with dithionite and ferricyanide...

---