Cloning and characterization of a maize cytochrome-b(5) reductase with Fe3+-chelate reduction capability

We previously purified an NADH-dependent Fe3+-chelate reductase (NFR) from maize roots with biochemical features of a cytochrome-b(5) reductase (b(5)R ) [Sparla, Bagnaresi, Scagliarini and Trost (1997) FEBS Lett. 414, 571-575] . We have now cloned a maize root cDNA that, on the basis of sequence infor mation, calculated parameters and functional assay, codes for NFR. Maize NF R has 66 % and 65 % similarity to mammal and yeast b(5)R respectively. It h as a deduced molecular mass of 31.17 kDa and a pi of 8.53. An uncharged reg ion is observed at its N-terminus but no myristoylation consensus site is p resent. Taken together, these results, coupled with previous biochemical ev idence, prove that NFR belongs to the b(5)R class and document b(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) clos ely resembles mammalian b(5)Rs. Several NFR isoforms are expected to exist in maize. They are probably not produced by alternative translational mecha nisms as occur in mammals, because of specific constraints observed in the maize NFR cDNA sequence. In contrast with yeast and mammals, tissue-specifi c and various subcellular localizations of maize b(5)R isoforms could resul t from differential expression of the various members of a multigene family . The first molecular characterization of a plant b(5)R indicates an overal l remarkable evolutionary conservation for these versatile reductase system s. In addition, the well-characterized Fe3+-chelate reduction capabilities of NFR, in addition to known Fe3+-haemoglobin reduction roles for mammal b( 5)R isoforms, suggest further and more generalized roles for the b(5)R clas s in endocellular iron reduction.