Identification and characterization of the propanediol utilization protein PduP of Lactobacillus reuteri for 3-hydroxypropionic acid production from glycerol |
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Authors: | Luo Lian Hua Seo Jeong-Woo Baek Jin-Oh Oh Baek-Rock Heo Sun-Yeon Hong Won-Kyung Kim Dae-Hyuk Kim Chul Ho |
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Institution: | (1) Microbe-based Fusion Technology Research Center, Jeonbuk Branch Institute, KRIBB, Jeongeup, Jeonbuk, 580-185, South Korea;(2) Institute for Molecular Biology and Genetics, Research Center of Bioactive Materials, Chonbuk National University, Jeonju, Chonbuk, 561-756, South Korea;(3) School of Life sciences and Biotechnology, Korea University, Seoul, 136-701, South Korea;(4) Interdisplinary Program of the Graduate School for Bioenergy & Biomaterials, Chonnam National University, Gwangju, 500-757, South Korea; |
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Abstract: | Although the de novo biosynthetic mechanism of 3-hydroxypropionic acid (3-HP) in glycerol-fermenting microorganisms is still
unclear, the propanediol utilization protein (PduP) of Lactobacillus species has been suggested to be a key enzyme in this regard. To verify this hypothesis, a pduP gene from Lactobacillus reuteri was cloned and expressed, and the encoded protein was characterized. Recombinant L. reuteri PduP exhibited broad substrate specificity including 3-hydroxypropionaldehyde and utilized both NAD+ and NADP+ as a cofactor. Among various aldehyde substrates tested, the specific activity was highest for propionaldehyde, at pH 7.8
and 37 °C. The K
m and V
max values for propionaldehyde in the presence of NAD+ were 1.18 mM and 0.35 U mg−1, respectively. When L. reuteri pduP was overexpressed in Klebsiella pneumoniae, 3-HP production remarkably increased as compared to the wild-type strain (from 0.18 g L−1 to 0.72 g L−1) under shake-flask culture conditions, and the highest titer (1.38 g L−1 3-HP) was produced by the recombinant strain under batch fermentation conditions in a bioreactor. This is the first report
stating the enzymatic properties of PduP protein and the probable role in biosynthesis of 3-HP in glycerol fermentation. |
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