Purification and properties of a nitrite reductase from porphyra yezoenis Ueda

概要

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Nitrite reductase was extracted from the red alga Porphyra yezoensis Ueda and purified through precipitation with ammonium sulfate, column chromatographies, and polyacrylamide gel disk electrophoresis. The enzyme prcparation thus obtained showed a single band on disk electrophoresis. The absorption spectrum had three maxima at 385 nm (Soret band), 580 nm (a-band), and 278 nm; the ratio of absorbance of the Soret band to the a-band was 4.3. The molecular weight and the number of amino acid residues were estimated to be 63,000 and 601, respectively. The enzyme activity was optimal at around pH 7.5, and its activity was heat labile as indicated by reduction of activity by about 70% when healed at 37℃ for 10 min. The enzyme used ferredoxin and methyl viologen, but not NADP^+ or NAD^+, as the electron carriers. Moreover, reduced forms of the latter two showed no effect on its activity. Km values of this enzyme for NO_2-, Fd, and MV were 8.1 × 10^<-4> M, 4.3 × 10^<-8> M, and 3.7 × 10^<-4> M, respectively. Almost half of its activity was lost when potassium cyanide was added at a concentration as low as 10^<-5> M, and the Ki value was 1.8 × 10^<-5> Thus, the nitrite reductase of Porphyra must be systematically grouped in EC 1.7.7.1. It resembled closely that of Chlorella, except for the amounts of some amino acids.
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