产蛋白酶芽孢杆菌的筛选鉴定及酶学特性分析

为了分离筛选产蛋白酶芽孢杆菌,以开发新型饲料添加剂,本研究选取小鼠肠道内容物为样品,通过对样品进行预处理以及酪蛋白平板检测法分离筛选获得产蛋白酶芽孢杆菌,对该菌进行形态学和分子生物学鉴定并分析其生长规律、产酶规律以及所产蛋白酶的酶学特性。结果表明,经形态学和分子生物学鉴定该菌为特基拉芽孢杆菌(Bacillus tequilensis),其所产蛋白酶的最适反应条件为50℃,pH=8.0,具有一定的高温耐受性,在弱酸、中性和弱碱性条件下表现出一定的稳定性,Ca²⁺和Mn²⁺能够显著提高酶活(P<0.05),Zn²⁺、Mg²⁺、Cu²⁺和EDTA、尿素溶液对蛋白酶活力具有显著抑制作用(P<0.05),培养至16 h菌体量达到最大值,发酵22 h上清液中的蛋白酶活力达到最高值(57.77 U/mL)。该芽孢杆菌的成功筛选分离以及其所产蛋白酶的分析研究,为新型饲料添加剂的开发利用提供了物质基础。

Screening,identification and characterization of protease-producing Bacillus

In order to isolate and screen the protease-producing Bacillus,and develop new feed additives,the intestinal contents of mice were obtained as samples,and the protease-producing Bacillus was isolated from the samples and was screened through pretreatment and casein plate detection.Then,the strain was identified by morphology and molecular biology,and its growth and enzyme production attributes and characteristics of the protease were analyzed.The results showed that the strain was identified as Bacillus tequilensis,and the optimal reaction condition for the protease was 50℃,pH 8.0,indicating that the protease had a certain thermostability,and it possessed certain stability under the conditions of weak acid,neutral and weak base,Ca²⁺and Mn²⁺increased the enzyme activity significantly(P<0.05).Zn²⁺,Mg²⁺,Cu²⁺,EDTA and urea solutions significantly inhibited the protease activity(P<0.05).When cultured for 16 h,the amount of bacteria reached the maximal value,and the protease activity in the supernatant reached the maximal value(57.77 U/mL)after 22 h fermentation.The successful screening and isolation of Bacillus in this study and the analysis of the protease produced by Bacillus provided materials and theoretical basis for development and utilization of new feed additives in the future.