海洋源蛋白酶产生菌筛选及酶学特性的初步研究

Preliminary Study on Screening of High Protease Production Strain from Marine and Its Enzymology Characteristics

从海洋源鱿鱼中筛选高产蛋白酶菌株。通过检测蛋白酶产生水解圈结合蛋白酶活性测定的方法筛选高产蛋白酶菌株,采用PCR技术对筛选菌株进行16S rRNA鉴定,并构建目的菌株的系统发育树,同时研究粗蛋白酶的酶学特性。结果表明:筛选得到的10株产蛋白酶活力较高的菌株,经鉴定分别属于芽孢杆菌属(Bacillus sp.)、普罗威登斯菌属(Providencia sp.)和假单胞菌属(Pseudomonas sp.)。其中SW5菌株产酶活性最高达257.67±2.44 U/mL,为甲基营养型芽孢杆菌(Bacillus methylotrophicus)。该菌株所产粗蛋白酶的酶学特性研究发现,其最适pH为8.0,最适温度为40℃,终离子浓度为1 mmol/L时Mn2+、Ba2+和Ca2+对该蛋白酶活性有较高的激活作用,而Fe^(2+)和Zn^(2+)能明显抑制该蛋白酶活性。

The purpose of this study is to screen high protease production strain from the squid,with generation protease hydrolysis circle and protease activity determination method. The typical strains are identified by 16 S rRNA sequencing,and subject to phylogenetic analysis. Meanwhile,the properties of crude protease enzyme are researched. The results show that after screening 10 strains had a high yield protease activity,identified belong to Bacillus sp.,Providencia sp.,Pseudomonas sp.,Among them SW5 strains produced highest enzyme activity up to 257.67±2.44 U/mL,identified as Bacillus methylotrophicus. Investigation on the crude enzymology characteristics further indicate that the enzyme exhibites the highest activity at pH 8.0 and the optimum temperature is 40 ℃. The protease activity is greatly inhibited by Mn2+、Ba2+and Ca2+,but is not inhibited by Fe2+and Zn2+at the final ion concentration 1 mmol/L.