高温蛋白酶产生菌的筛选及其产酶条件和酶学性质分析

Screening of Thermostable Protease-producing Bacterium and Analysis of Fermentation Conditions and Protease Properties

从徂徕山温泉附近土样中分离到9株产高温蛋白酶菌株,选取一株碱性蛋白酶高产菌株L7为出发菌株,进行显微形态、16S rRNA基因序列分析,将其初步鉴定为短芽孢杆菌(Brevibacillus sp.)。研究该菌株发酵条件,确定产酶的最佳培养基组成为葡萄糖40 g/L,蛋白胨20 g/L,磷酸氢二钠1.4 g/L,氯化钙0.6 g/L,硫酸镁0.4 g/L,通过培养基优化,酶活达到103.08 U/mL。最佳培养条件为250 mL三角烧瓶中装液量50 mL、pH8.0、培养温度为55℃、培养时间为24 h。对该菌株酶学性质研究,L7菌株所产高温蛋白酶的最适温度为55℃,最适pH为10,并且具有良好的温度稳定性和pH稳定性,酶活性受PMSF强烈抑制。

Nine thermophilic bacterial strains with thermophilic protease activity were collected from the soil around a hot spring in Mountain Culai.The strain which showed high protease activity was selected for protease assay and named as L7.Based on morphological characteristics and 16S rRNA gene sequence analysis,strain L7 was identified as Brevibacillus sp..The optimal medium components were glucose 40 g/L,peptone 20 g/L,Na2HPO4 1.4 g/L,CaCl2 0.6 g/L,MgCl2 0.4 g/L.The protease activity can achieve 103.08 U/mL through the medium optimization.The optimal culture conditions were 50 mL medium in 250 mL Erlenmeyer flask,initial pH8.0,at 55℃for 24 h.Enzymatic properties showed that the enzyme optimal temperature is 55℃,optimum pH10.0.The enzyme exhibited high thermal stability and stable at a wide range of pH,and its activity is intensively const rained by PMSF.