经硫酸铵分级沉淀、DEAE-Cellulose柱层析、SephadexG-75凝胶过滤层析,由巨大芽孢杆菌(B.megaterium MPF-906)分离纯化得到亚硝酸还原酶(nitrite reductase,NiR),分子量为35ku.NiR反应的最适温度和pH值分别为40℃和6.5.热稳定性较好,80℃保温4h后仍有50％的酶活力.在pH5.5～9.0均较稳定,残余酶活都在60％以上.Cu2+、Ba2+能提高酶活力;Mn2+、Pb2+对酶活力有明显抑制作用;Na+、Fe3+、M2+、Al3+对其有轻微抑制作用；Ca2+、Zn2+对酶活力影响不大.亚硝酸钠为底物,该酶的Km=8.6mmol/L,Vmax=4.1U/mg,该酶的适电子供体为抗坏血酸20mol/L、0.1mol/L的连二亚硫酸钠和0.075mol/L的草酸钠.%The nitrite reductase (NiR) from Bacillus megaterium MPF-906 was isolated and purified by ammonium sulfate salting-out, DEAE-cellu-lose column chromatography and SephadexG-75 gel filtration, and the molecular weight was 35ku. The optimal temperature and pH value for enzyme reaction were 40"C and 6.5, respectively. The enzyme has good thermal stability, which remained about 50% enzyme activity after treatment at 80t for 4h. It was in stable state when pH value in between of 5.5 and 9.0 and the activity of residual enzyme was above 60%. Besides, activity of NiR was strongly inhibited by Mn2*, Pb2* and moderately inhibited by Na*, Fe**, Mg2*, AP, while Ca2*, Zn2* had no evident effects. Its Km and V?, were 8.6mmol/L and 4.1U/mg, respectively when using sodium nitrite as substrate, and the optimal electronic donor was 20mmol/L ascorbic acid, O.lmoML sodium hydrosulfite and 0.075mol/L sodium oxalate.
展开▼
