摘要
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+、Mg2+、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-cellulose column chromatography and SephadexG-75 gel filtration, and the molecular weight was 35ku. The optimal temperature and pH value for enzyme reaction were 40℃ and 6.5, respectively. The enzyme has good thermal stability, which remained about 50% enzyme activity after treatment at 80℃ 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 Mn^2＋, Pb^2＋ and moderately inhibited by Na^＋, Fe^3＋, Mg^2＋, Al^3＋, while Ca^2＋, Zn^2＋ had no evident effects. Its Km and Vmax were 8.6mmol/L and 4.1U/mg, respectively when using sodium nitrite as substrate, and the optimal electronic donor was 20mmoUL ascorbic acid, 0. I mol/L sodium hydrosulfite and 0.075mol/L sodium oxalate.
出处
《中国酿造》
CAS
北大核心
2011年第9期22-27,共6页
China Brewing
基金
国家863计划课题(2008AA10Z306)
国家自然基金(20876164)
关键词
巨大芽孢杆菌
亚硝酸还原酶
分离纯化
酶学性质
Bacillus megaterium
nitrite reductase
isolation and purification
enzyme properties
