Crystal structure of the FAD-containing fragment of corn nitrate reductase at 2.5 A resolution: relationship to other flavoprotein reductases

Abstract

Background: In the biological assimilation of nitrate in plants and microorganisms, nitrate is reduced to ammonium by transfer of eight electrons in a two-step process. The first step of the pathway, the reduction of nitrate to nitrite, is catalyzed by nitrate reductase, a multi-redox cofactor enzyme which belongs to the class of flavoprotein pyridine nucleotide cytochrome reductases. The enzyme can be divided into three functional fragments that bind the cofactors molybdopterin, heme-iron and flavin adenine dinucleotide (FAD)/nicotinamide adenine dinucleotide (NADH).

Results: Here we describe the crystal structure of the recombinant cytochrome b reductase fragment of corn nitrate reductase, in complex with the cofactor FAD, determined to 2.5 A resolution. This catalytically competent fragment of nitrate reductase consists of two domains, the amino-terminal lobe, which binds FAD, and the carboxy-terminal lobe, which presumably binds NADH, connected by a linker region.

Conclusions: Nitrate reductase belongs to the class of flavoprotein pyridine nucleotide cytochrome reductases, a subgroup in the family of ferredoxin reductase-like flavoproteins. Comparison with other members of this family reveals that large structural differences are found in the relative orientation of the cofactor binding lobes. This indicates that conformational changes might be important for biological function.