Chlorella nitrate reductase

Cannons, A.C., Iida, N. Solomonson, L.P. (1991). Expression of a cDNA clone encoding the haem-binding domain of Chlorella nitrate reductase. Biochemical Journal 278, 203-9. 

Zeiler, K.G. Solomonson, L.P. (1989). Regulation of Chlorella nitrate reductase control of enzyme activity and immunoreactive protein levels by ammonia. Archives of Biochemistry and Biophysics 269, 46-54. 

Reduction of Cu and Fe organic complexes is saturable, and is inhibited by cell-impermeable probes, including polyclonal antibodies raised to Chlorella nitrate reductase (Jones and Morel, 1987). The model proposed suggests that a fraction of a membrane-bound nitrate reductase (the so-called diaphorase component that catalyzes NADH reduction), spans the cell membrane (Fig. 4). Organic metal complexes intercept electrons (destined for nitrate reduction intracellularly) at the outer cell surface. 

The conversion in vitro of the active form of Chlorella nitrate reductase into the inactive form depends on its reduction by NAD(P)H, and is reversible by reoxidation with ferri-cyanide. Inactivation by NADH requires the first moiety of the complex to be active and proceeds much faster at high pH or when ADP at low concentration (0 3 mM) is simultaneously present. This synergistic effect is quite specific for NADH and ADP. Nitiate, as well as several of its competitive inhibitors, completely prevents and even reverses inactivation by NADH and ADP. In fact, Vennesland and co-workers have demonstrated the presence of cyanide in the in vivo inactivated enzyme by overnight incubation with nitrate and phosphate. ... 

Chlorella vulgaris nitrate reductase structure, 3,1438 Chlorides... 

Study by X-ray absorption spectroscopy of the extended X-ray absorption fine structure (EXAFS) has provided estimates of both the nature and the number of the nearest neighboring atoms around the Mo. The EXAFS spectra of xanthine dehydrogenase and of nitrate reductase from Chlorella confirmed the... 

Nitrate reductase from Chlorella, an assimilatory enzyme, is a homotetramer of molecular weight 360 000 and contains one each of Mo, heme and FAD per subunit. The nitrate reductase from E. coli is a dissimilatory enzyme. EXAFS data are available on the molybdenum sites in both enzymes (Table 24).1050 The environment of the molybdenum in the assimilatory enzyme is similar to that found for sulfite oxidase, with at least two sulfur ligands near the molybdenum and a shuttle between monoxo and dioxo forms with redox change in the enzyme. This allows a similar mechanism to be put forward for the assimilatory nitrate reductase,1051 shown in equation (57), where an oxo group is transferred from nitrate to MoIV with production of nitrite and MoVI. 

The assimilatory nitrate reductase from Chlorella contains the molybdenum cofactor, as evidenced by the ability of the enzyme to donate the cofactor to the nitrate reductase of the mutant nit-1 of N. crassa. Reduction of the enzyme with NADH gives the Mov ESR signal, which is abolished on reoxidation with nitrate. Line shape and g values of the signal show a pH dependence similar to those observed previously for sulfite oxidase. The signal observed at pH 7.0 shows evidence for interaction with a single exchangeable proton.1053... 

Both assimilatory and dissimilatory nitrate reductases are molybdoenzymes, which bind nitrate at the molybdenum. EXAFS studies1050 have shown that there are structural differences between the assimilatory nitrate reductase from Chlorella vulgaris and the dissimilatory enzyme from E. coli. The Chlorella enzyme strongly resembles sulfite oxidase1050,1053 and shuttles between mon-and di-oxo forms, suggesting an oxo-transfer mechanism for reduction of nitrate. This does not appear to be the case for the E. coli enzyme, for which an oxo-transfer mechanism seems to be unlikely. The E. coli enzyme probably involves an electron transfer and protonation mechanism for the reduction of nitrate.1056 It is noteworthy that the EXAFS study on the E. coli nitrate reductase showed a long-distance interaction with what could be an electron-transfer subunit. 

Barber, M.J. Solomonson, L.P. (1986). The role of the essential sulfhydryl group in assimilatory NADH nitrate reductase of Chlorella. Journal of Biological Chemistry 261, 4562-7. 

Stohr, C., Schuler, F., and Tischner, R., 1995, Glycosyl-phosphatidylinositol-anchored proteins exist in the plasma membranes of Chlorella Saccharophila (Kruger) Nadson Plasma-membrane-bound nitrate reductase as an example. Planta 196 284-287. 

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). 

Tischner, R. A., Ward, M. R., and Huffaker, R. C. (1989). Evidence for a plasma-membrane-bound nitrate reductase involved in nitrate uptake of Chlorella sorokiniana. Planta. 178, 19—24. 

Nitrate reductase (assimilatory) Chlorella vulgaris 360000 4 4 heme, 4FAD... 

Cerium tetrakis(acetylacetonate), 1114 Cesium complexes crown ethers, 40 Chevrel phases, 1321 Chlorella vulgaris nitrate reductase structure, 1438 Chlorine cations... 

Pistorius et al. (1978) have indicated that the time course kinetics of accumulation of cyanide inactivated enzyme observed, when cells of Chlorella are transferred from nitrate to ammonium, are too slow to account for the measured rapid decline in nitrate reduction. They suggest that cessation of nitrate utilization when this organism is transferred to ammonium is due to a termination of nitrate uptake. However, Diez et al. (1977) indicated that in Ankistrodesmus the absence of a nitrogen source (i.e., no nitrate uptake) brings about a much smaller inactivation of nitrate reductase than that produced by the transfer from nitrate to ammonium. Thus, the rapid in vivo inactivation of nitrate reduction observed in this alga when it is transferred to... 

The single cell green alga Chlorella produces a nitrate reductase that has been investigated by Kay et al The natural reductant is NADPH and in the presence of NR and NADPH the reduced NR enzyme is able to donate electrons to cytochrome c as evident from the appearance of a catalytic cytochrome c oxidation wave. Reduced MV and diquat were effective artificial electron donors to NR and in the presence of nitrate a catalytic reduction current was observed. 

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