Cloning and characterization of a maize cytochrome-b sub(5) reductase with Fe super(3+)-chelate reduction capability

We previously purified an NADH-dependent Fe super(3+)-chelate reductase (NFR) from maize roots with biochemical features of a cytochrome-b sub(5) reductase (b sub(5)R). We have now cloned a maize root cDNA that, on the basis of sequence information, calculated parameters and functional assay, codes for NFR. Maize NFR has 66% and 65% similarity to mammal and yeast b sub(5)R respectively. It has a deduced molecular mass of 31.17 kDa and a pI of 8.53. An uncharged region is observed at its N-terminus but no myristoylation consensus site is present. Taken together, these results, coupled with previous biochemical evidence, prove that NFR belongs to the b sub(5)R class and document b sub(5)R from a plant at the molecular level for the first time. We have also identified a putative Arabidopsis thaliana NFR gene. Its organization (nine exons) closely resembles mammalian b sub(5)Rs. Several NFR isoforms are expected to exist in maize. They are probably not produced by alternative translational mechanisms as occur in mammals, because of specific constraints observed in the maize NFR cDNA sequence. In contrast with yeast and mammals, tissue-specific and various subcellular localizations of maize b sub(5)R isoforms could result from differential expression of the various members of a multigene family. The first molecular characterization of a plant b sub(5)R indicates an overall remarkable evolutionary conservation for these versatile reductase systems. In addition, the well-characterized Fe super(3+)-chelate reduction capabilities of NFR, in addition to known Fe super(3+)-haemoglobin reduction roles for mammal b sub(5)R isoforms, suggest further and more generalized roles for the b sub(5)R class in endocellular iron reduction.