Abstract
Objective
Glycerol-based biosynthesis of 3-hydroxypropionic acid (3-HP) in Klebsiella pneumoniae involves two reactions: glycerol conversion to 3-hydroxypropionaldehyde (3-HPA) by glycerol dehydratase, and 3-HPA conversion to 3-HP by aldehyde dehydrogenase (ALDH). The ALDH catalysis consumes a lot of cofactor nicotinamide adenine dinucleotide (NAD+), which constrains 3-HP production.
Results
Here we report that intensifying niacin-based biosynthesis of NAD+ can substantially enhance 3-HP production. We constructed tac promoter-driven NAD+ synthesis pathway in K. pneumoniae. The strain only overexpressing nicotinate phosphoribosyltransferase (PncB) showed 14.24% increase in the production of NAD+ relative to the stain harboring an empty vector. When PncB was coexpressed with PuuC (one of native ALDHs), the recombinant strain exhibited increased ALDH activity but slightly reduced 3-HP production due to plasmid burden. When 30 mg niacin l−1 (a substrate for biosynthesis of NAD+) was added into shake flask, the strain produced 0.55 g 3-HP l−1, which was 2.75 times that of the control. In a 5-L bioreactor, replenishment of niacin led to 36.43% increase of 3-HP production.
Conclusions
These results indicated that intensifying niacin-based biosynthesis of NAD+ boosts 3-HP production.
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References
Ansede JH, Pellechia PJ, Yoch DC (1999) Metabolism of acrylate to β-hydroxypropionate and its role in dimethylsulfoniopropionate lyase induction by a salt marsh sediment bacterium, Alcaligenes faecalis M3A. Appl Environ Microbiol 65:5075–5081
Ashok S, Raj SM, Rathnasingh C, Park S (2011) Development of recombinant Klebsiella pneumoniae ∆dhaT strain for the co-production of 3-hydroxypropionic acid and 1,3-propanediol from glycerol. Appl Microbiol Biotechnol 90:1253–1265. https://doi.org/10.1007/s00253-011-3148-z
Ashok S, Sankaranarayanan M, Ko Y, Jae KE, Ainala SK, Kumar V, Park S (2013) Production of 3-hydroxypropionic acid from glycerol by recombinant Klebsiella pneumoniae∆dhaT∆yqhD which can produce vitamin B12 naturally. Biotechnol Bioeng 110:511–524. https://doi.org/10.1002/bit.24726
Black WB, Zhang L, Mak WS, Maxel S, Cui Y, King E, Fong B, Sanchez Martinez A, Siegel JB, Li H (2020) Engineering a nicotinamide mononucleotide redox cofactor system for biocatalysis. Nat Chem Biol 16:87–94. https://doi.org/10.1038/s41589-019-0402-7
Celinska E (2010) Debottlenecking the 1,3-propanediol pathway by metabolic engineering. Biotechnol Adv 28:519–530. https://doi.org/10.1016/j.biotechadv.2010.03.003
Croft T, James Theoga Raj C, Salemi M, Phinney BS, Lin SJ (2018) A functional link between NAD+ homeostasis and N-terminal protein acetylation in Saccharomyces cerevisiae. J Biol Chem 293:2927–2938. https://doi.org/10.1074/jbc.M117.807214
Dong WR, Sun CC, Zhu G, Hu SH, Xiang LX, Shao JZ (2014) New function for Escherichia coli xanthosine phophorylase (xapA): genetic and biochemical evidences on its participation in NAD+ salvage from nicotinamide. BMC Microbiol 14:29. https://doi.org/10.1186/1471-2180-14-29
Forage RG, Foster MA (1982) Glycerol fermentation in Klebsiella pneumoniae: functions of the coenzyme B12-dependent glycerol and diol dehydratases. J Bacteriol 149:413–419
Huang Y, Li Z, Shimizu K, Ye Q (2013) Co-production of 3-hydroxypropionic acid and 1,3-propanediol by Klebseilla pneumoniae expressing aldH under microaerobic conditions. Bioresour Technol 128:505–512. https://doi.org/10.1016/j.biortech.2012.10.143
Ji D, Wang L, Hou S, Liu W, Wang J, Wang Q, Zhao ZK (2011) Creation of bioorthogonal redox systems depending on nicotinamide flucytosine dinucleotide. J Am Chem Soc 133:20857–20862. https://doi.org/10.1021/ja2074032
Knaus T, Paul CE, Levy CW, De Vries S, Mutti FG, Hollmann F, Scrutton NS (2016) Better than nature: nicotinamide biomimetics that outperform natural coenzymes. J Am Chem Soc 138:1033–1039. https://doi.org/10.1021/jacs.5b12252
Kumar V, Park S (2018) Potential and limitations of Klebsiella pneumoniae as a microbial cell factory utilizing glycerol as the carbon source. Biotechnol Adv 36:150–167. https://doi.org/10.1016/j.biotechadv.2017.10.004
Li Y, Su MY, Ge XZ, Tian PF (2013) Enhanced aldehyde dehydrogenase activity by regenerating NAD+ in Klebsiella pneumoniae and implications for the glycerol dissimilation pathways. Biotechnol Lett 35:1609–1615. https://doi.org/10.1007/s10529-013-1243-1
Li Y, Wang X, Ge X, Tian P (2016) High production of 3-hydroxypropionic acid in Klebsiella pneumoniae by systematic optimization of glycerol metabolism. Sci Rep 6:26932. https://doi.org/10.1038/srep26932
Liang L, Liu R, Wang G, Gou D, Ma J, Chen K, Jiang M, Wei P, Ouyang P (2012) Regulation of NAD(H) pool and NADH/NAD+ ratio by overexpression of nicotinic acid phosphoribosyltransferase for succinic acid production in Escherichia coli NZN111. Enzyme Microb Technol 51:286–293. https://doi.org/10.1016/j.enzmictec.2012.07.011
Liu Y, Feng Y, Wang L, Guo X, Liu W, Li Q, Wang X, Xue S, Zhao ZK (2019) Structural insights into phosphite dehydrogenase variants favoring a non-natural redox cofactor. ACS Catal 9:1883–1887. https://doi.org/10.1021/acscatal.8b04822
Matsakas L, Hrůzová K, Rova U, Christakopoulos P (2018) Biological production of 3-hydroxypropionic acid: an update on the current status. Fermentation 4:13. https://doi.org/10.3390/fermentation4010013
Mohan Raj S, Rathnasingh C, Jung WC, Park S (2009) Effect of process parameters on 3-hydroxypropionic acid production from glycerol using a recombinant Escherichia coli. Appl Microbiol Biotechnol 84:649–657. https://doi.org/10.1007/s00253-009-1986-8
Niu K, Cheng XL, Qin HB, Liu JS, Zheng YG (2017) Investigation of the key factors on 3-hydroxypropionic acid production with different recombinant strains. 3 Biotech 7:314. https://doi.org/10.1007/s13205-017-0966-4
Paul CE, Gargiulo S, Opperman DJ, Lavandera I, Gotor-Fernandez V, Gotor V, Taglieber A, Arends IWCE, Hollmann F (2013) Mimicking nature: synthetic nicotinamide cofactors for C=C bioreduction using enoate reductases. Org Lett 15:180–183. https://doi.org/10.1021/ol303240a
Schwarz M, Kopcke B, Weber RW, Sterner O, Anke H (2004) 3-Hydroxypropionic acid as a nematicidal principle in endophytic fungi. Phytochemistry 65:2239–2245. https://doi.org/10.1016/j.phytochem.2004.06.035
Talarico TL, Casas IA, Chung TC, Dobrogosz WJ (1988) Production and isolation of reuterin, a growth inhibitor produced by Lactobacillus reuteri. Antimicrob Agents Chemother 32:1854–1858. https://doi.org/10.1128/aac.32.12.1854
Tokuyama K, Ohno S, Yoshikawa K, Hirasawa T, Tanaka S, Furusawa C, Shimizu H (2014) Increased 3-hydroxypropionic acid production from glycerol, by modification of central metabolism in Escherichia coli. Microb Cell Fact 13:64. https://doi.org/10.1186/1475-2859-13-64
Wang L, Ji D, Liu Y, Wang Q, Wang X, Zhou YJ, Zhang Y, Liu W, Zhao ZK (2017a) Synthetic cofactor-linked metabolic circuits for selective energy transfer. ACS Catal 7:1977–1983. https://doi.org/10.1021/acscatal.6b03579
Wang L, Liu B, Liu Y, Sun Y, Liu W, Yu D, Zhao ZK (2019) Escherichia coli strain designed for characterizing in vivo functions of nicotinamide adenine dinucleotide analogues. Org lett 21:3218–3222. https://doi.org/10.1021/acs.orglett.9b00935
Wang X, Zhou YJ, Wang L, Liu W, Liu Y, Peng C, Zhao ZK (2017b) Engineering Escherichia coli nicotinic acid mononucleotide adenylyltransferase for fully active amidated NAD biosynthesis. Appl Environ Microbiol 83:e00692–e717. https://doi.org/10.1128/AEM.00692-17
Zhang Q, Gong JS, Dong TT, Liu TT, Li H, Dou WF, Lu ZM, Shi JS, Xu ZH (2017) Nitrile-hydrolyzing enzyme from Meyerozyma guilliermondii and its potential in biosynthesis of 3-hydroxypropionic acid. Bioprocess Biosyst Eng 40:901–910. https://doi.org/10.1007/s00449-017-1754-6
Acknowledgements
This study was funded by grants from National Key Research and Development Program of China (2018YFA0901800), National Natural Science Foundation of China (21476011) and National High Technology Research and Development Program (863 Program) (2015AA021003).
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Wu, S., Zhao, P., Li, Q. et al. Intensifying niacin-based biosynthesis of NAD+ to enhance 3-hydroxypropionic acid production in Klebsiella pneumoniae. Biotechnol Lett 43, 223–234 (2021). https://doi.org/10.1007/s10529-020-03011-y
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DOI: https://doi.org/10.1007/s10529-020-03011-y