细胞破碎方法对Bacillus sp. DL-2胞内蛋白酶拆分乙酸苏合香酯的影响

doi:10.11978/2020041

热带海洋学报 ›› 2021, Vol. 40 ›› Issue (2): 39-48.doi: 10.11978/2020041CSTR: 32234.14.2020041

细胞破碎方法对Bacillus sp. DL-2胞内蛋白酶拆分乙酸苏合香酯的影响

王晓敏^1,⁴(), 董璐^1,⁴, 张继福³, 张云^1,², 胡云峰^1,²()

1.中国科学院南海海洋研究所, 中国科学院热带海洋生物资源与生态重点实验室, 广东省海洋药物重点实验室, 广东广州 510301
2.南方海洋科学与工程广东省实验室(广州), 广东广州 511458
3.广东省中医院, 广东广州 510120
4.中国科学院大学, 北京 100049

收稿日期:2020-04-15 修回日期:2020-05-12 出版日期:2021-03-10 发布日期:2020-06-01
通讯作者: 胡云峰
作者简介:王晓敏(1995—), 山东省烟台市人, 硕士研究生, 主要从事酶催化研究。email: wangxiaomin18@mails.ucas.ac.cn
基金资助:
南方海洋科学与工程广东省实验室(广州)人才团队引进重大专项(GML2019ZD0406);广东省自然科学基金项目(2018A030313151);广东省海洋渔业科技攻关与研发方向项目(A201701C12);中国科学院战略性先导科技专项(XDA11030404);中国科学院“科学”号高端用户项目(KEXUE2018G05)

The effects of different cell breaking methods on the asymmetrical hydrolysis of phenylethyl acetate using intracellular proteases of Bacillus sp. DL-2

WANG Xiaomin^1,⁴(), DONG Lu^1,⁴, ZHANG Jifu³, ZHANG Yun^1,², HU Yunfeng^1,²()

1. CAS Key Laboratory of Tropical Marine Bio-resources and Ecology, Guangdong Key Laboratory of Marine Materia Medical, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, China
2. Southern Marine Science and Engineering Guangdong Laboratory (Guangzhou), Guangzhou 511458, China
3. Guangdong Provincial Hospital of Chinese Medicine, Guangzhou 510120, China
4. University of Chinese Academy of Sciences, Beijing 100049, China

Received:2020-04-15 Revised:2020-05-12 Online:2021-03-10 Published:2020-06-01
Contact: HU Yunfeng
Supported by:
Key Special Project for Introduced Talents Team of Southern Marine Science and Engineering Guangdong Laboratory(Guangzhou)(GML2019ZD0406);Natural Science Foundation of Guangdong Province of China(2018A030313151);Scientific and Technological Project of the Ocean and Fishery from Guangdong Province of China(A201701C12);Priority Research Program of the Chinese Academy of Sciences(XDA11030404);Senior User Project of the Research Vessel KEXUE(KEXUE2018G05)

摘要/Abstract

摘要：

(R)-1-苯乙醇是手性药物合成的关键手性砌块, 在多种手性化合物的合成及医药、香料的工业生产中具有重要的作用。对筛选自西太平洋深海沉积物的Bacillus sp. DL-2进行破碎, 获取的胞内蛋白酶能有效拆分(±)-乙酸苏合香酯, 制备高光学纯的(R)-1-苯乙醇。为使拆分效果最优化, 本试验对比了表面活性剂、溶菌酶和超声法3种不同的破碎细胞方式, 并对表面活性剂和溶菌酶破碎法进行了单因素条件优化。最优方法及优化的反应条件为: 使用体积分数0.5%的表面活性剂曲拉通X-100在35℃和pH7.0条件下破碎细胞2h。经过优化后, (R)-1-苯乙醇的对映体过量值为96%, 转化率为23%, 为利用胞内蛋白酶生产手性化学品提供了参考。

关键词: 表面活性剂, 溶菌酶, 超声破碎, 手性拆分, (R)-1-苯乙醇

Abstract:

(R)-1-phenylethanol is a key chiral building block in the synthesis of chiral drugs, and plays important roles in the synthesis of many chiral compounds and in the industrial production of medicines and spices. The strain Bacillus sp. DL-2 screened from the deep-sea sediments of the western Pacific Ocean was broke, and the obtained intracellular proteases can asymmetrically hydrolyze (±)-1-phenylethyl acetate to obtain (R)-1-phenylethanol. Three different ways, namely, surfactant, lysozyme and ultrasonic, were utilized to crush Bacillus sp. DL-2. After the optimizations of the surfactant and lysozyme crushing methods, the optimal working conditions were found to be 0.5% TritonX-100, 35 ℃ and pH 7.0 for 2 h. After process optimization, (R)-1-phenylethanol was obtained with an enantiomeric excess of 96% and a conversion of 23%, which provide reference for using intracellular proteases to produce chiral chemicals.

Key words: surfactant, lysozyme, ultrasonication, kinetic resolution, (R)-1-phenylethanol

中图分类号:

P735.4

王晓敏, 董璐, 张继福, 张云, 胡云峰. 细胞破碎方法对Bacillus sp. DL-2胞内蛋白酶拆分乙酸苏合香酯的影响[J]. 热带海洋学报, 2021, 40(2): 39-48.

WANG Xiaomin, DONG Lu, ZHANG Jifu, ZHANG Yun, HU Yunfeng. The effects of different cell breaking methods on the asymmetrical hydrolysis of phenylethyl acetate using intracellular proteases of Bacillus sp. DL-2[J]. Journal of Tropical Oceanography, 2021, 40(2): 39-48.

图/表 5

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图5

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细胞破碎方法对Bacillus sp. DL-2胞内蛋白酶拆分乙酸苏合香酯的影响

The effects of different cell breaking methods on the asymmetrical hydrolysis of phenylethyl acetate using intracellular proteases of Bacillus sp. DL-2

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