原核表达重组牛凝乳酶原及重组牛凝乳酶酶学特性  

Prokaryotic expression of the recombinant bovine prochymosin and analysis on enzymatic properties of the recombinant chymosin

作　　者：普燕[1] 李轶杰[1] 张富春[1] 

机构地区：[1]新疆生物资源基因工程重点实验室,新疆大学生命科学与技术学院,新疆乌鲁木齐830046

出　　处：《食品与发酵工业》2013年第8期13-19,共7页Food and Fermentation Industries

基　　金：新疆自治区动物学重点学科-乳品工程资助项目(2011001)

摘　　要：摘要凝乳酶能够专一性裂解κ-酪蛋白，是制造干酪的关键酶。运用大肠杆菌表达系统对牛凝乳酶原进行了原核表达和初步纯化，活化重组凝乳酶原及测定凝乳活性，对重组凝乳酶的酶学特性进行分析。结果表明i大肠杆菌表达的重组蛋白约占菌体总蛋白的66．3％，每升培养液可纯化约200mg的重组凝乳酶原，活化后的凝乳酶活力可达600000SU／g。经测定凝乳酶最适作用温度为57～62℃，并在pH2～7、低于40℃的温度范围内稳定。金属离子中Al3＋，Fe3＋和Cu2＋能显著增强酶活；胃蛋白酶抑制剂pepstatinA对酶有明显的抑制作用。Chymosin is the key enzyme that can specifically cleave K-casein and manufacture cheese. In this study, we expressed the bovine proehymosin in Escherichia coli, purified and activated the recombinant prochymosin, measured its milk-clotting activity, then analyzed the enzymatic properties of chymosin. The results showed that the prokaryotic expressed prochymosin accounted for about 66.3% of the total bacterial protein, the purified prochymosin was about 200 mg/L of culture liquid, and chymosin activity was 600 000 SU/g after proehymosin activation. The op- timal temperature range of milk-clotting activity of recombinant chymosin was 57 - 62 ℃ , and the chymosin was rela- tively stable in pH 2 - 7 and below 40℃. Metal ions such as Al3＋ , Fe3＋ and Cu2＋ significantly increased the activity of chymosin, however, pepsin inhibitor pepstatin A had an obvious inhibitory effect on the activity of chymosin. It would provide a high expression strain for the industrial production of ehymosin.

关 键 词：牛凝乳酶原 凝乳酶 原核表达 凝乳酶活力 酶学特性 

分 类 号：TS252.1[轻工技术与工程—农产品加工及贮藏工程]