An Efficient Prephenate Dehydrogenase Gene for the Biosynthesis of L-tyrosine: Gene Mining, Sequence Analysis, and Expression Optimization

Abstract

L-tyrosine is a key precursor for synthesis of various functional substances, but the microbial production of L-tyrosine faces huge challenges. The development of new microbial chassis cell and gene resource is especially important for the biosynthesis of L-tyrosine. In this study, the optimal host strain Bacillus amyloliquefaciens HZ-12 was firstly selected by detecting the production capacity of L-tyrosine. Subsequently, the recombinant expression of 15 prephenate dehydrogenase genes led to the discovery of the best gene, Bao-tyrA from B. amyloliquefaciens HZ-12. After the overexpression of Bao-tyrA, the L-tyrosine yield of the recombinant strain HZ/P43-Bao-tyrA reach 411 mg/L, increased by 42% compared with the control strain (HZ/pHY300PLK). Moreover, the nucleic acid sequence and deduced amino acid sequence of the gene Bao-tyrA were analyzed, and their conservative sites and catalytic mechanisms were proposed. Finally, the expression of Bao-tyrA was regulated through a promoter and 5′-UTR sequence to obtain the optimal expression elements. Thereby, the maximum L-tyrosine yield of 475 mg/L was obtained from HZ/P43-UTR3-Bao-tyrA. B. amyloliquefaciens was applied for the first time to produce L-tyrosine, and the optimal prephenate dehydrogenase gene Bao-tyrA and corresponding expression elements were obtained. This study provides new microbial host and gene resource for the construction of efficient L-tyrosine chassis cells, and also lays a solid foundation for the production of various functional tyrosine derivatives.

Cloning Systems in Bacillus: Bioengineering of Metabolic Pathways for Valuable Recombinant Products

“…P 43 was compared to 13 other promoters, including one dual (P 43 -P ylB ), of different strengths: eight of them it surpassed in a statistically significant way; while there was no significant difference with the other five, P 43 was nevertheless able to confer greater production of L-tyrosine than any of them (about 400 mg/L). Five different UTRs were also tested, but only one produced a significant difference, and then only 16% (475 mg/L) [151].…”