Activation of Nitrite Reductase in a Cell-Free System from the Denitrifier Alcaligenes sp. by Freeze-Thawing

概要

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Nitrite reductase (NiR) activity of the cell-free extract or the soluble fraction prepared from cells of Alcaligenes sp. NCIB 11015 grown anaerobically in the presence of nitrate was examined by measuring the rate of nitrite disappearance with dithionitemethyl viologen (MV) as an electron donor. Freezing at -20℃ and subsequent thawing of the fraction resulted in 5-40 times increase of the specific activity of NiR. From the experiments on the effect of freezing conditions on the activation, the phase change of solvent water due to freezing is considered to play an important role in the activation. This activation occurred with the preparation in the exponential growth phase, but not that in the stationary growth phase. Clearly, the low-molecular-weight (< 12,000) component which was obtained from the soluble fraction through a collodion bag participated in the activation. The activated enzyme proved to be the dissimilatory NiR, because NO production from nitrite, one of the typical characteristics of the dissimilatory NiR, was also activated when assayed with ascorbate-tetramethyl-p-phenylene diamine (TMPD) as an electron donor. Nevertheless, the reaction products of nitrite reduction were identified as hydroxylamine and ammonia with dithionite-MV. The possible pathway of nitrite reduction with this electron donor is discussed.
日本植物生理学会の論文