摘要
以本实验室构建的能高效表达海参i-型溶菌酶的毕赤酵母基因工程菌HS 3-1为出发菌株,利用5 L发酵罐进行扩大培养,发酵液经分离纯化,精制得到海参i-型溶菌酶。采用MALDI-TOF质谱分析,确定了该溶菌酶的相对分子质量为14709。进一步对酶的生化特性进行检测,结果表明其最适温度为35℃,最适pH为6.5,该溶菌酶在25~80℃范围内活性保持在80%以上,说明其具有良好的热稳定性。分子动力学模拟结果表明,在热处理过程中,溶菌酶蛋白的结构域中出现一段“H1-L2-H2”的构象,并且其活性位点位于两个螺旋区域(H1和H2)上,推测这是该溶菌酶分子能够保持高度的热稳定性的主要原因。
The recombinant strain Pichia pastoris HS 3-1 was used to express i-type lysozyme of sea cucumber efficiently.The type i-type lysozyme was isolated and purified from the fermentation broth by 5 L fermentor.The molecular weight of the lysozyme was determined to be 14709 by MALDI-TOF mass spectrometry.The results showed that the optimal temperature and pH were 35℃and 6.5,respectively.The activity of lysozyme remained above 80%in the range of 25-80℃,indicating that the lysozyme had good thermal stability.Molecular dynamics simulation results showed that a“H1-L2-H2”conformation appeared in the domain of the protein during heat treatment,and its active sites located in two helix regions(H1 and H2).It is maybe the main reason for the high thermal stability of the lysozyme.
作者
马锁
丛丽娜
谢定刚
陈清平
MA Suo;CONG Lina;XIE Dinggang;CHEN Qingping(School of Biological Engineering, Dalian Polytechnic University, Dalian 116034, China)
出处
《大连工业大学学报》
CAS
北大核心
2020年第3期174-178,共5页
Journal of Dalian Polytechnic University
基金
辽宁省自然科学基金项目(201602047)。
关键词
海参
溶菌酶
酶学性质
分子动力学
sea cucumber
lysozyme
enzymatic properties
molecular dynamics
