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. 1983;2(11):1909–1914. doi: 10.1002/j.1460-2075.1983.tb01678.x

On the presence of a heme-binding domain homologous to cytochrome b5 in Neurospora crassa assimilatory nitrate reductase

Kim Hô Diêp Lê 1, Florence Lederer 1,*
PMCID: PMC555387  PMID: 16453480

Abstract

Assimilatory nitrate reductase has been purified with 55% recovery from a Neurospora crassa nmr-1 nit-6 mutant, using a modification of a published procedure. It possesses one heme per 240 000 g, and subunits of mol. wt. 68 000. Upon digestion with chymotrypsin, a heme-binding domain was isolated by gel filtration; its visible spectrum was highly similar to that of cytochrome b5. On SDS gels, the fraction showed two heme-containing bands of ˜10 000 and 12 5000 daltons; their amino acid composition was not very different, suggesting that they originated from the same region of the polypeptide chain. After S-carboxymethylation, the mixture of bands was submitted to cyanogen bromide cleavage, and the fragments were separated by h.p.l.c. The two largest fragments yielded an identical sequence upon automated degradation. This sequence (39 residues with some gaps) could be easily aligned with that of cytochrome b5 starting close to the N terminus. These results are discussed in terms of the possible quaternary structure of N. crassa nitrate reductase, whose heme-binding domain proves to be another member of the family of b5-like cytochromes.

Keywords: cytochrome b5, evolution, Neurospora crassa, nitrate reductase, structural domain

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Selected References

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