Monday, April 19, 2010
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Production of 3-hydroxypropionaldehyde from glycerol in recombinant Escherichia coli expressing novel glycerol dehydratase gene cluster from Lactobacillus brevis

Su-Ryang Kwak1, Kwang-Wook Kim1, Jin-Ah Noh1, Yong-Cheol Park2, and Jin-Ho Seo1. (1) Department of Agricultural Biotechnology, Seoul National University, Room 200A-Bio8104 San 56-1, Shillim-dong, Kwanak-gu, Seoul, 151-742, South Korea, (2) Department of Advanced Fermentation Fusion Science and Technology, Kookmin University, 861-1 Jeongneung-Dong Seongbux-Gu, Seoul, South Korea

3-Hydroxypropionaldehyde (3-HPA) is an important intermediate for production of value-added chemicals of 1,3-propanediol and 3-hydroxypropionic acid. Glycerol dehydratase catalyzes the reaction of glycerol dehydration to produce 3-HPA. In this study, the dhaB gene cluster coding for a novel glycerol dehydratase was cloned from Lactobacillus brevis ATCC 367 and expressed in recombinant Escherichia coli for production of 3-HPA from glycerol. The L. brevis dhaB gene cluster is composed of three open reading frames of dhaB1, dhaB2 and dhaB3. The deduced proteins of DhaB1, DhaB2 and DhaB3 have the theoretical molecular weights of 61,411, 25,658 and 19,361 Da, respectively, of which amino acid sequences showed high similarity to other glycerol dehydratases of L. reuteri and L. collinoides. For the expression of the L. brevis dhaB cluster in recombinant E. coli, two translational start codons of TTG and GTG were replaced by ATG and the dhaB gene cluster was designed to be located at the downstream of the IPTG inducible T7 promoter. Optimum reaction conditions for L. brevis glycerol dehydratase were determined at pH 7.0 and 20 μM coenzyme B12. Meanwhile, two hypothetical genes of dhaR1 and dhaR2 showed moderate homologies to the genes encoding glycerol dehydratase reactivases and cloned from L. brevis. Recombinant E. coli BL21 star (DE3)/pELDRR expressing the dhaB gene cluster and dhaR gene cluster produced 0.30 g/L 3-HPA at 23% yield in a batch fermentation with LB medium containing 4.40 g/L glycerol.

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