桔青霉产核酸酶P1酶分离纯化及其酶学性质

桔青霉（Penicillium citrinum）产核酸酶P1浓缩液采用活性炭脱色、硫酸铵分级沉淀、脱盐和凝胶层析等分离技术，得到核酸酶P1纯组分，并研究了该酶的酶学性质。该酶纯化后比酶活达到33967 U/mg，纯化倍数为8.48倍；该酶的米氏常数K_m、最大反应速度V_m和催化常数K_cat分别为2.50 mmol/L、0.0864 mmol/（mL·min）和252.43 s?1。该酶最适温度为75 ℃，热稳定范围60～75 ℃；最适pH为5.5，pH稳定范围为4.0～6.0；Zn2+在1 mmol/L条件下对核酸酶P1有很好的激活作用，Cu2+和Co2+对该酶的抑制作用明显，而Ni2+、Fe2+、Mn2+等离子具有不同程度的抑制作用。本研究对于该酶的广泛应用奠定了科学基础。

Isolation,Purification and Enzymatic Properties of Nuclease P1 Fermented by Penicillium citrinum

The nuclease P1 was purified to obtain pure component by activated carbon decolorization,(NH4)2 SO4 precipitation, desalination and gel chromatography and its enzymatic properties was investigated. This purified enzyme had a specific activity of 33967 U/mg protein after 8.48-fold purification. The Michaelis constant(Km), the maximum reaction rates(Vm) and the catalytic constant(Kcat) of the purified enzyme were 2.50 mmol/L, 0.0864 mmol/(mL·min) and 252.43 s-1,respectively. The optimization pH and temperature for the nuclease P1 were at pH5.5 and 75 ℃. The enzyme was stable in the temperature range from 60 to 75 ℃ and in the pH range from 4.0 to 6.0. Zn2+ had a positive effect on the enzyme activity, while Cu2+ was a strong inhibitor of nuclease P1, Ni2+、Fe2+、Mn2+ had the different inhibition. This research laid a scientific foundation for the extensive application of the enzyme.