Enhancement of pipecolic acid production by the expression of multiple lysine cyclodeaminase in the Escherichia coli whole-cell system
Yeong-Hoon Han, Tae-Rim Choi, Ye-Lim Park, Jun Young Park, Hun-Suk Song, Hyun-Joong Kim, Sun Mi Lee, Sol Lee Park, Hye Soo Lee, Shashi Kant Bhatia, Ranjit Gurav, Yung-Hun Yang*
Department of Biological Engineering, College of Engineering, Konkuk University, 1 Hwayang-dong, Gwangjin-gu, Seoul 05029, Republic of Korea
*Corresponding author (E-mail:seokor@konkuk.ac.kr )
Running title: Reinforced-LCD improves pipecolic acid production
ABSTRACT
Pipecolic acid, a non-proteinogenic amino acid, is a metabolite in lysine metabolism and a key chiral precursor in local anesthesia and macrolide antibiotics. To replace the environmentally unfriendly chemical production or preparation procedure of pipecolic acid, many biological synthetic routes have been studied for a long time. Among them, synthesis by lysine cyclodeaminase (LCD), encoded by pipA , has several advantages, including stability of enzyme activity and NAD+ self-regeneration. Thus, we selected this enzyme for pipecolic acid biosynthesis in a whole-cell bioconversion. To construct a robust pipecolic acid production system, we investigated important conditions including expression vector, strain, culture conditions, and other reaction parameters. The most important factors were introduction of multiple pipA genes into the whole-cell system and control of agitation. As a result, we produced 724 mM pipecolic acid (72.4% conversion), and the productivity was 0.78 g/L/h from 1 M l-lysine after 5 days. This is the highest production reported to date.
Keywords : pipecolic acid (l-PA), pharmaceutical precursor, whole-cell bioconversion, lysine cyclodeaminse (LCD), reinforced enzyme