一株海洋细菌Pseudomonas sp.7～11所产蛋白酶的纯化及部分性质研究  

Studies on purification and characterization of a protease from a marine bacterium, Pseudomons sp. 7~11

作　　者：刘清海[1] 荆谷[1] 冯静[1] 孔健[1] 马桂荣[1] 

机构地区：[1]山东大学微生物技术国家重点实验室,济南250100

出　　处：《中国海洋药物》2001年第3期20-24,共5页Chinese Journal of Marine Drugs

摘　　要：海洋假单胞菌 (Pseudomonassp.7～ 11)菌株所产蛋白酶经盐析、透析、离子交换层析后 ,得到了聚丙烯酰胺凝胶电泳纯的一酶组分。该酶的比活力从 19.4 6 U· mg-1提高到 13953115U· mg-1,提高了 717.0 2倍 ,回收率为 1.2 4 %。酶水解酪蛋白的最适作用温度为 4 5℃ ,最适 p H为 8.0 ;酶在 p H6～ 7,50℃以下时比较稳定。 EDTA和 IAA强烈抑制酶的活性 ,SDS也具有一定的抑制作用 ,而 PMSF对该酶的抑制作用不明显 ,酶被 EDTA失活后 ,Mn2 +、Mg2 +能部分恢复其活力 ,尤其是 Mn2 ;Hg2 +、Fe2 +、Zn2 +、Cu2 +和 Pb2 +对酶有抑制作用 ,而 Ca2 +、Mg2 +和 (NH4 ) 2 SO4 则有一定的激活作用 ;该酶对乙醇和尿素有较强的抗性 ,对吐温 2 0也具有一定的抗性。纯酶的相对分子质量大约 30 0 0 0A protease from the culture supernatant of marine bacteria Pseudomonas sp. 7~11 was discovered. One component of the protease was purified to homogeneity by ammonium sulfate precipitation, dialysis, ion exchange chromatography. The specific activity of the enzyme was raised from 19.46U·mg -1 to 13953115U·mg -1 ,which was 717.02 times that of the culture supernatant with a yield of 1.24%.The molecular mass of purified enzyme was estimated to be 3000Da about by SDS PAGE. The optimum temperature for the activity was observed to be 45℃ using casein as substrate.The optimum pH for activity the enzyme was 8.0 and it was stable between pH6～7 and below 50℃.The activity of the enzyme was inhibited by EDTA and IAA strongly. But was not inhibited by PMSF. the enzyme was inhibited Mn 2+ , Hg 2+ , Fe 2+ , Zn 2+ , Cu 2+ ,Pb 2+ and SDS. The inactivity of the enzyme with EDTA could be recovered partially by Mg 2+ .while Ca 2+ , Mg 2+ and (NH 4) 2SO 4 was activator for the activity of the enzyme. The activity of the enzyme was fairly stable in the presence of ethanol and urea, and was resistant to Tween 20.

关 键 词：蛋白酶 海洋假单胞菌 纯化 酶性质 

分 类 号：R282.77[医药卫生—中药学] R284[医药卫生—中医学]