Nitrate reductase

Prokaryotic nitrate reductases belong to the DMSO reductase family of molybdoenzymes and have been classified into three groups, assimilatory nitrate reductases (Nas), respiratory nitrate reductase (Nar), and periplasmic nitrate reductases (Nap). The active site of these enzymes is a Mo ion that is bound to the four thiolate functions of two pterin molecules. The coordination sphere of the Mo is completed by one amino-acid side chain and oxygen and/or sulfur ligands. The exact environment of the Mo ion in certain of these enzymes (oxygen versus sulfur as a sixth molybdenum ligand) is still debated. The Mo is covalently attached to the protein by a cysteine ligand in Nap, and an aspartate in Nar.^[2]

Structure

The transmembrane respiratory nitrate reductase (EC) is composed of three subunits; an alpha, a beta and two gamma. It is the second nitrate reductase enzyme which it can substitute for the NRA enzyme in Escherichia coli allowing it to use nitrate as an electron acceptor during anaerobic respiration.^[3]

Nitrate reductase gamma subunit resembles cytochrome b and transfers electrons from quinones to the beta subunit.^[4]

The nitrate reductase of higher plants is a cytosolic protein. There exists a GPI-anchored variant that is found on the outer face of the plasma membrane. Its exact function is still not clear.^[5]

A transmembrane nitrate reductase that can function as a proton pump (similar to the case of anaerobic respiration) has been discovered in a diatom Thalassiosira weissflogii.^[6]

Catalytic mechanism

Nitrate molecule binds to the active site with the Mo ion in the +6 oxidation state. Electron transfer to the active site occurs only in the proton-electron transfer stage, where the MoV species plays an important role in catalysis. The presence of the sulfur atom in the molybdenum coordination sphere creates a pseudo-dithiolene ligand that protects it from any direct attack from the solvent. Upon the nitrate binding there is a conformational rearrangement of this ring that allows the direct contact of the nitrate with MoVI ion. This rearrangement is stabilized by the conserved methionines Met141 and Met308. The reduction of nitrate into nitrite occurs in the second step of the mechanism where the two dimethyl-dithiolene ligands have a key role in spreading the excess of negative charge near the Mo atom to make it available for the chemical reaction. The reaction involves the oxidation of the sulfur atoms and not of the molybdenum as previously suggested. The mechanism involves a molybdenum and sulfur-based redox chemistry instead of the currently accepted redox chemistry based only on the Mo ion. The second part of the mechanism involves two protonation steps that are promoted by the presence of MoV species. MoVI intermediates might also be present in this stage depending on the availability of protons and electrons. Once the water molecule is generated only the MoVI species allow water molecule dissociation, and, the concomitant enzymatic turnover.^[7]

Applications

Nitrate reductase activity can be used as a biochemical tool for predicting grain yield and grain protein production.^[8]^[9]

Nitrate reductase promotes amino acid production in tea leaves.^[10] Under south Indian conditions, it is reported that tea plants sprayed with various micronutrients (like Zn, Mn and B) along with Mo enhanced the amino acid content of tea shoots and also the crop yield.^[11]

References

↑ PDB: 1Q16; Bertero MG, Rothery RA, Palak M, Hou C, Lim D, Blasco F, Weiner JH, Strynadka NC (September 2003). "Insights into the respiratory electron transfer pathway from the structure of nitrate reductase A". Nat. Struct. Biol. 10 (9): 681–7. doi:10.1038/nsb969. PMID 12910261.
↑ Tavares P, Pereira AS, Moura JJ, Moura I (December 2006). "Metalloenzymes of the denitrification pathway". J. Inorg. Biochem. 100 (12): 2087–100. doi:10.1016/j.jinorgbio.2006.09.003. PMID 17070915.
↑ Blasco F, Iobbi C, Ratouchniak J, Bonnefoy V, Chippaux M (June 1990). "Nitrate reductases of Escherichia coli: sequence of the second nitrate reductase and comparison with that encoded by the narGHJI operon". Mol. Gen. Genet. 222 (1): 104–11. doi:10.1007/BF00283030. PMID 2233673.
↑ Pantel I, Lindgren PE, Neubauer H, Götz F (July 1998). "Identification and characterization of the Staphylococcus carnosus nitrate reductase operon". Mol. Gen. Genet. 259 (1): 105–14. doi:10.1007/s004380050794. PMID 9738886.
↑ Tischner R (October 2000). "Nitrate uptake and reduction in higher and lower plants". Plant, Cell and Environment 23 (10): 1005–1024. doi:10.1046/j.1365-3040.2000.00595.x.
↑ Jones GJ, Morel FM (May 1988). "Plasmalemma redox activity in the diatom thalassiosira: a possible role for nitrate reductase". Plant Physiol. 87 (1): 143–7. doi:10.1104/pp.87.1.143. PMC 1054714. PMID 16666090.
↑ Cerqueira NM, Gonzalez PJ, Brondino CD, Romão MJ, Romão CC, Moura I, Moura JJ (November 2009). "The effect of the sixth sulfur ligand in the catalytic mechanism of periplasmic nitrate reductase". J Comput Chem 30 (15): 2466–84. doi:10.1002/jcc.21280. PMID 19360810.
↑ Croy LI, Hageman RH (1970). "Relationship of nitrate reductase activity to grain protein production in wheat". Crop Science 10 (3): 280–285. doi:10.2135/cropsci1970.0011183X001000030021x.
↑ Dalling MJ, Loyn RH (1977). "Level of activity of nitrate reductase at the seedling stage as a predictor of grain nitrogen yield in wheat (Triticum aestivum L.)". Australian Journal of Agricultural Research 28 (1): 1–4. doi:10.1071/AR9770001.
↑ Ruan J, Wu X, Ye Y, Härdter R (1988). "Effect of potassium, magnesium and sulphur applied in different forms of fertilisers on free amino acid content in leaves of tea (Camellia sinensis L". J. Sci. Food Agric. 76 (3): 389–396. doi:10.1002/(SICI)1097-0010(199803)76:3<389::AID-JSFA963>3.0.CO;2-X.
↑ Venkatesan S (November 2005). "Impact of genotype and micronutrient applications on nitrate reductase activity of tea leaves". J. Sci. Food Agric. 85 (3): 513–516. doi:10.1002/jsfa.1986.

External links

UMich Orientation of Proteins in Membranes protein/pdbid-1q16
Nitrate reductases at the US National Library of Medicine Medical Subject Headings (MeSH)

Oxidoreductases: nitrogenous donor (EC 1.7)

1.7.1	GMP reductase

1.7.2	Nitrite reductase (NO-forming)

1.7.3	Urate oxidase

1.7.7	Ferredoxin—nitrite reductase

1.7.99	Hydroxylamine reductase

Enzymes

Activity	Active site Binding site Catalytic triad Oxyanion hole Enzyme promiscuity Catalytically perfect enzyme Coenzyme Cofactor Enzyme catalysis Enzyme kinetics Lineweaver–Burk plot Michaelis–Menten kinetics

Regulation	Allosteric regulation Cooperativity Enzyme inhibitor

Classification	EC number Enzyme superfamily Enzyme family List of enzymes

Types	EC1 Oxidoreductases(list) EC2 Transferases(list) EC3 Hydrolases(list) EC4 Lyases(list) EC5 Isomerases(list) EC6 Ligases(list)

Protein: flavoproteins

Acetolactate synthase Acyl CoA dehydrogenase Apoptosis-inducing factor Butyryl CoA dehydrogenase Cryptochrome Cytochrome b5 reductase Cytokinin dehydrogenase Dihydrolipoamide dehydrogenase Flavodoxin Methemoglobin reductase Methylenetetrahydrofolate reductase NADH dehydrogenase NADPH oxidase Nitrate reductase Sarcosine oxidase Thioredoxin reductase

This article incorporates text from the public domain Pfam and InterPro IPR003816

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