利用酪氨酸酚裂解酶通过全细胞生物转化法生产 L-酪氨酸。

Production of L-tyrosine using tyrosine phenol-lyase by whole cell biotransformation approach.

机构信息

Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China; National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, China.

Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China.

出版信息

Enzyme Microb Technol. 2019 Dec;131:109430. doi: 10.1016/j.enzmictec.2019.109430. Epub 2019 Sep 12.

DOI:10.1016/j.enzmictec.2019.109430

PMID:31615664

Abstract

L-tyrosine is an amino acid that has been widely used in the food, agriculture and pharmaceutical industries. In order to screen a tyrosine phenol-lyase (TPL) with excellent catalytic performance for L-tyrosine production, TPL genes from Citrobacter freundii (CfTPL), Erwinia herbicola (EhTPL) and Rhodobacter capsulatus (TutA) were codon-optimized and overexpressed in Escherichia coli. The results showed that EhTPL had the highest whole cell catalysis activity and tyrosine yield (3-fold that of CfTPL). The results of RT-qPCR and a stability analysis also revealed that EhTPL had a higher transcriptional level in whole cell catalysis, while CfTPL possessed greater stability. Conditions for the production by whole cell transformation were optimized in terms of reaction conditions and fed-batch strategy. Finally, the maximum production was obtained with a titer of 48.5 g·L by intermittent feeding with a conversion ratio of 75%.

摘要

L-酪氨酸是一种氨基酸，广泛应用于食品、农业和制药行业。为了筛选出一种用于生产 L-酪氨酸的具有优异催化性能的酪氨酸酚裂解酶（TPL），我们对柠檬酸杆菌（CfTPL）、草生欧文氏菌（EhTPL）和荚膜红假单胞菌（TutA）的 TPL 基因进行了密码子优化，并在大肠杆菌中过表达。结果表明，EhTPL 具有最高的全细胞催化活性和酪氨酸产率（是 CfTPL 的 3 倍）。RT-qPCR 分析和稳定性分析的结果还表明，EhTPL 在全细胞催化过程中有更高的转录水平，而 CfTPL 则具有更高的稳定性。我们还从反应条件和补料分批策略两个方面对全细胞转化生产的条件进行了优化。最终，通过间歇补料的方式，获得了 48.5 g·L-1 的最高产量，转化率为 75%。