Effects of freezing on purified nitrite reductase from a denitrifier, Alcaligenes sp. NCIB 11015.
The effects of freezing on Alcaligenes sp. nitrite reductase [nitric-oxide: ferricytochrome c oxidoreductase, EC 1.7.2.1] dissolved in sodium phosphate (pH 7.2) were investigated. The nitrite reductase was gradually activated with time in the frozen state, resulting in an increase in its activity of 2.5-4.5 times. The final freezing temperature influenced the enzyme activation, maximal activation being observed at around -20 degrees C. All the enzymatic activities that the nitrite reductase is known to catalyze were enhanced by freeze-thawing. The activation was followed by neither association-dissociation nor any gross conformational change of the enzyme molecule, but was accompanied by an increase in the fluorescence intensity of 2-p-toluidinonaphthalene-6-sulfonate used as a hydrophobic probe. The results are consistent with the hypothesis that the activation of the NiR is due to a limited conformational change of the enzyme molecule, particularly in the hydrophobic region. The mechanism of the activation of NiR by freeze-thawing is discussed, in comparison with the mechanisms of inactivation by freeze-thawing of many enzymes reported by previous workers.
D000421:Alcaligenes / Q000201:enzymology*; D001426:Bacterial Proteins / Q000032:analysis; D002850:Chromatography, Gel; D002942:Circular Dichroism; D004789:Enzyme Activation; D005615:Freezing; D009247:NADH, NADPH Oxidoreductases / Q000032:analysis*; D009572:Nitrite Reductases / Q000032:analysis*; D011489:Protein Denaturation; D013997:Time Factors
D001426:Bacterial Proteins; D009247:NADH, NADPH Oxidoreductases; D009572:Nitrite Reductases