摘要
为了解枯草芽孢杆菌JNB001发酵所产蛋白酶的特性,试验以蛋白酶活力为指标,研究了pH、温度等因素对酶促反应的影响,并通过水解度考察了粗酶液对各种动物蛋白的酶解效果。结果表明,酶反应最适pH为8.0,温度为45℃;粗酶液在pH 5.5~9.0的范围内较稳定,过碱则不利于酶液的保存。在45℃下热稳定性良好,超过50℃保温10min后残余酶活已不足60%;金属离子Ca^(2+)和Mg^(2+)、还原剂DTT、表面活性剂Tween-80等能大幅度提高酶活力。而Fe^(2+)、Cu^(2+)、金属螯合剂EDTA、变性剂SDS等物质则会抑制蛋白酶活性;酶促反应的Km为6.43mg/mL,Vmax为47.39μg/min;针对不同蛋白底物,酶促反应较适pH为7.0~9.0,温度为40~50℃。该粗酶液具有水解猪毛、猪蹄甲、脱毛牛皮、明胶等硬蛋白类物质的能力。上述试验结果可为枯草芽孢杆菌JNB001在病死畜禽无害化生物处理体系等方面的工业应用提供参考。
This experiment was aimed to study the protease properties produced by Bacillus subtilis JNB001 via the index of protease activity to investigate the effects of pH,temperature and other factors on the enzymatic reaction,and the ability of the crude enzyme solution to hydrolyze various animal protein was measured by the degree of hydrolysis.The results showed that the most suitable pH of reaction was 8.0and the temperature was 45℃.The crude enzyme solution in the range of pH 5.5to 9.0was relatively stable,while it was not suitable for the preservation of the enzyme solution when pH was over 9.0.At 45℃it had good thermal stability when exceed 50℃for 10 min,the residual activity had been less than 60%.Metal ions such as Ca^2＋and Mg^2＋,reducing agent like DTT,surfactant as Tween-80,etc.could greatly improve the enzyme activity.On the opposite,Fe^2＋and Cu^2＋as well as other ions,metal chelator EDTA,denaturant SDS and other materials could inhibit the protease activity.The Km was 6.43 mg/mL,and Vmaxwas47.39μg/min.It had a range of reaction pH from 7.0to 9.0and temperature from 40 to 50℃in the situation for different protein substrates.The crude enzyme solution could hydrolyze pig hair,pig nail,hair removal of cowhide,gelatin and other hard protein substances.The results provided the strong date support for the Bacillus subtilis JNB001 which could be used in dead livestock harmless biological treatment system and other aspects of industrial applications.
出处
《中国畜牧兽医》
CAS
北大核心
2015年第12期3261-3267,共7页
China Animal Husbandry & Veterinary Medicine
基金
宁波市科技计划重大专项(2013C11033)
关键词
蛋白酶
酶学性质
动物蛋白
酶解效果
protease
enzyme properties
animal protein
enzymatic hydrolysis effect
