Engineering of Baeyer-Villiger monooxygenase-based Escherichia coli biocatalyst for large scale biotransformation of ricinoleic acid into (Z)-11-(heptanoyloxy)undec-9-enoic acid.
Baeyer-Villiger monooxygenases (BVMOs) are able to catalyze regiospecific Baeyer-Villiger oxygenation of a variety of cyclic and linear ketones to generate the corresponding lactones and esters, respectively. However, the enzymes are usually difficult to express in a functional form in microbial cells and are rather unstable under process conditions hindering their large-scale applications. Thereby, we investigated engineering of the BVMO from Pseudomonas putida KT2440 and the gene expression system to improve its activity and stability for large-scale biotransformation of ricinoleic acid (1) into the ester (i.e., (Z)-11-(heptanoyloxy)undec-9-enoic acid) (3), which can be hydrolyzed into 11-hydroxyundec-9-enoic acid (5) (i.e., a precursor of polyamide-11) and n-heptanoic acid (4). The polyionic tag-based fusion engineering of the BVMO and the use of a synthetic promoter for constitutive enzyme expression allowed the recombinant Escherichia coli expressing the BVMO and the secondary alcohol dehydrogenase of Micrococcus luteus to produce the ester (3) to 85 mM (26.6 g/L) within 5 h. The 5 L scale biotransformation process was then successfully scaled up to a 70 L bioreactor; 3 was produced to over 70 mM (21.9 g/L) in the culture medium 6 h after biotransformation. This study demonstrated that the BVMO-based whole-cell reactions can be applied for large-scale biotransformations.
Department of Food Science and Engineering, Ewha Womans University, Seoul 120-750, Republic of Korea.
Joo-Hyun
JH
Seo
Department of Food Science and Engineering, Ewha Womans University, Seoul 120-750, Republic of Korea.
Hwan-Hee
HH
Kim
Department of Food Science and Engineering, Ewha Womans University, Seoul 120-750, Republic of Korea.
Eun-Yeong
EY
Jeon
AP Technology, Suwon, Kyunggi 443-702, Republic of Korea.
Young-Ha
YH
Song
AP Technology, Suwon, Kyunggi 443-702, Republic of Korea.
Chul-Soo
CS
Shin
Department of Food Science and Engineering, Ewha Womans University, Seoul 120-750, Republic of Korea.
Jin-Byung
JB
Park
D019149:Bioreactors / Q000382:microbiology*; D001711:Biotransformation / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism; D016958:Pseudomonas putida / Q000201:enzymology / Q000235:genetics; D012277:Ricinoleic Acids / Q000737:chemistry*
D012277:Ricinoleic Acids; D006899:Mixed Function Oxygenases; C030521:ricinoleic acid