Abstract
Sporolactobacillus inulinus is a superior d-lactic acid-producing bacterium and proposed species for industrial production. The major pathway for d-lactic acid biosynthesis, glycolysis, is mainly regulated via the two irreversible steps catalyzed by the allosteric enzymes, phosphofructokinase (PFK) and pyruvate kinase. The activity level of PFK was significantly consistent with the cell growth and d-lactic acid production, indicating its vital role in control and regulation of glycolysis. In this study, the ATP-dependent PFK from S. inulinus was expressed in Escherichia coli and purified to homogeneity. The PFK was allosterically activated by both GDP and ADP and inhibited by phosphoenolpyruvate; the addition of activators could partly relieve the inhibition by phosphoenolpyruvate. Furthermore, monovalent cations could enhance the activity, and Na+ was the most efficient one. Considering this kind activation, NaOH was investigated as the neutralizer instead of the traditional neutralizer CaCO3. In the early growth stage, the significant accelerated glucose consumption was achieved in the NaOH case probably for the enhanced activity of Na+-activated PFK. Using NaOH as the neutralizer at pH 6.5, the fermentation time was greatly shortened about 22 h; simultaneously, the glucose consumption rate and the d-lactic acid productivity were increased by 34 and 17%, respectively. This probably contributed to the increased pH and Na+-promoted activity of PFK. Thus, fermentations by S. inulinus using the NaOH neutralizer provide a green and highly efficient d-lactic acid production with easy subsequent purification.
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References
Andersen HW, Solem C, Hammer K, Jensen PR (2001) Twofold reduction of phosphofructokinase activity in Lactococcus lactis results in strong decreases in growth rate and in glycolytic flux. J Bacteriol 183:3458–3467
Bai ZZ, Gao Z, He BF, Wu B (2015) Effect of lignocellulose-derived inhibitors on the growth and D-lactic acid production of Sporolactobacillus inulinus YBS1-5. Bioprocess Biosyst Eng 38:1993–2001
Bai ZZ, Gao Z, Sun JD, Wu B, He BF (2016) D-Lactic acid production by Sporolactobacillus inulinus YBS1-5 with simultaneous utilization of cottonseed meal and corncob residue. Bioresour Technol 207:346–352
Belouski E, Watson DE, Bennett GN (1998) Cloning, sequence, and expression of the phosphofructokinase gene of Clostridium acetobutylicum ATCC 824 in Escherichia coli. Curr Microbiol 37:17–22
Bennett BD, Kimball EH, Gao M, Osterhout R, Van Dien SJ, Rabinowitz JD (2009) Absolute metabolite concentrations and implied enzyme active site occupancy in Escherichia coli. Nat Chem Biol 5:593–599
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254
Branny P, de La Torre F, Garel JR (1996) The genes for phosphofructokinase and pyruvate kinase of Lactobacillus delbrueckii subsp. bulgaricus constitute an operon. J Bacteriol 178:4727–4230
Byrnes M, Zhu X, Younathan ES, Chang SH (1994) Kinetic characteristics of phosphofructokinase from Bacillus stearothermophilus: MgATP nonallosterically inhibits the enzyme. Biochemistry 33:3424–3431
Datta R, Henry M (2006) Lactic acid: recent advances in products, processes and technologies—a review. J Chem Technol Biot 81:1119–1129
Ding ZJ, Bai ZZ, Sun ZH, Ouyang PK, He BF (2004) Study on fermentative production of D(-)-lactic acid from glucose by Sporolactobacillus sp. Chin J Bioprocess Eng 2:30–36
Fothergill-Gilmore LA, Michels PA (1993) Evolution of glycolysis. Prog Biophys Mol Biol 59:105–235
Gao C, Ma CQ, Xu P (2011) Biotechnological routes based on lactic acid production from biomass. Biotechnol Adv 29:930–939
Gohara DW, Di Cera E (2016) Molecular mechanisms of enzyme activation by monovalent cations. J Biol Chem. doi:10.1074/jbc.R116.737833
Gong YH, Li TY, Li SY, Jiang ZY, Yang Y, Huang JL, Liu ZB, Sun HX (2016) Achieving high yield of lactic acid for antimicrobial characterization in cephalosporin resistance Lactobacillus by the co-expression of the phosphofructokinase and glucokinase. J Microbiol Biotechnol 26:1148–1161
Le Bras G, Deville-Bonne D, Garel JR (1991) Purification and properties of the phosphofructokinase from Lactobacillus bulgaricus. A non-allosteric analog of the enzyme from Escherichia coli. Eur J Biochem 198:683–687
Llanos RM, Harris CJ, Hillier AJ, Davidson BE (1993) Identification of a novel operon in Lactococcus lactis encoding three enzymes for lactic acid synthesis: phosphofructokinase, pyruvate kinase, and lactate dehydrogenase. J Bacteriol 175:2541–2551
Ludwig H, Homuth G, Schmalisch M, Dyka FM, Hecker M, Stülke J (2001) Transcription of glycolytic genes and operons in Bacillus subtilis: evidence for the presence of multiple levels of control of the gapA operon. Mol Microbiol 41:409–422
Mosser R, Reddy MCM, Bruning JB, Sacchettini JC, Reinhart GD (2012) Structure of the Apo form of Bacillus stearothermophilus phosphofructokinase. Biochemistry 51:769–775
Nakano S, Ugwu CU, Tokiwa Y (2012) Efficient production of D-(-)-lactic acid from broken rice by Lactobacillus delbrueckii using Ca(OH)2 as a neutralizing agent. Bioresour Technol 104:791–794
Paricharttanakul NM, Ye S, Menefee AL, Javid-Majd F, Sacchettini JC, Reinhart GD (2005) Kinetic and structural characterization of phosphofructokinase from Lactobacillus bulgaricus. Biochemistry 44:15280–15286
Poorman RA, Randolph A, Kemp RG, Heinrikson RL (1984) Evolution of phosphofructokinase—gene duplication and creation of new effector sites. Nature 309:467–469
Qin JY, Wang XW, Zheng ZJ, Ma CQ, Tang HZ, Xu P (2010) Production of L-lactic acid by a thermophilic Bacillus mutant using sodium hydroxide as neutralizing agent. Bioresour Technol 101:7570–7576
Riley-Lovingshimer MR, Reinhart GD (2005) Examination of MgATP binding in a tryptophan-shift mutant of phosphofructokinase from Bacillus stearothermophilus. Arch Biochem Biophys 436:178–186
Riley-Lovingshimer MR, Ronning DR, Sacchettini JC, Reinhart GD (2002) Reversible ligand-induced dissociation of a tryptophan-shift mutant of phosphofructokinase from Bacillus stearothermophilus. Biochemistry 41:12967–12974
Shirakihara Y, Evans PR (1988) Crystal structure of the complex of phosphofructokinase from Escherichia coli with its reaction products. J Mol Biol 204:973–994
Sun J, Wang Y, Wu B, Bai Z, He B (2015) Enhanced production of D-lactic acid by Sporolactobacillus sp.Y2-8 mutant generated by atmospheric and room temperature plasma. Biotechnol Appl Biochem 62:287–292
Tlapak-Simmons VL, Reinhart GD (1994) Comparison of the inhibition by phospho(enol)pyruvate and phosphoglycolate of phosphofructokinase from B. stearothermophilus. Arch Biochem Biophys 308:226–230
Tsuji H (2005) Poly(lactide) stereocomplexes: formation, structure, properties, degradation, and applications. Macromol Biosci 5:569–597
Valdez BC, French BA, Younathan ES, Chang SH (1989) Site-directed mutagenesis in Bacillus stearothermophilus fructose-6-phosphate 1-kinase. Mutation at the substrate-binding site affects allosteric behavior. J Biol Chem 264:131–135
Xu TT, Bai ZZ, Wang LJ, He BF (2010) Breeding of D (–)-lactic acid high producing strain by low-energy ion implantation and preliminary analysis of related metabolism. Appl Biochem Biotechnol 160:314–321
Yon J, Fried M (1989) Precise gene fusion by PCR. Nucleic Acids Res 17:4895
Yu B, Su F, Wang LM, Xu K, Zhao B, Xu P (2011) Draft genome sequence of Sporolactobacillus inulinus strain CASD, an efficient D-lactic acid-producing bacterium with high-concentration lactate tolerance capability. J Bacteriol 193:5864–5865
Zhang DR, Zheng L, Wu B, He BF (2016) Characterization of D-lactate-dehydrogenase isozymes from a D-lactic acid-producing bacterium Sporolactobacillus inulinus. Acta Microbiol Sin. doi:10.13343/j.cnki.wsxb.20160133
Zhao B, Wang L, Li F, Hua D, Ma C, Ma Y, Xu P (2010) Kinetics of D-lactic acid production by Sporolactobacillus sp. strain CASD using repeated batch fermentation. Bioresour Technol 101:6499–6505
Zheng L, Bai ZZ, Xu TT, He BF (2012) Glucokinase contributes to glucose phosphorylation in D-lactic acid production by Sporolactobacillus inulinus Y2-8. J Ind Microbiol Biotechnol 39:1685–1692
Zheng L, Bai ZZ, Xu TT, He BF (2014a) Cloning and expression of phosphofructokinase gene from Sporolactobacillus inulinus in Escherichia coli. Chin J Bioprocess Eng 12:37–42
Zheng L, Xu TT, Bai ZZ, He BF (2014b) Mn2+/Mg2+-dependent pyruvate kinase from a D-lactic acid-producing bacterium Sporolactobacillus inulinus: characterization of a novel Mn2+-mediated allosterically regulated enzyme. Appl Microbiol Biotechnol 98:1583–1593
Zhu LF, Xu XL, Wang LM, Dong H, Yu B, Ma YH (2015) NADP+-preferring D-lactate dehydrogenase from Sporolactobacillus inulinus. Appl Environ Microbiol 81:6294–6301
Acknowledgements
This study was funded by grants from the National High Technology Program Research and Development Program of China (2012AA022200), the National Science Foundation of China (41401284), the Natural Science Foundation of Jiangsu (BK20141456), and the Field Frontier Program of the Institute of Soil Science (ISSASIP1640).
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Zheng, L., Liu, M., Sun, J. et al. Sodium ions activated phosphofructokinase leading to enhanced d-lactic acid production by Sporolactobacillus inulinus using sodium hydroxide as a neutralizing agent. Appl Microbiol Biotechnol 101, 3677–3687 (2017). https://doi.org/10.1007/s00253-017-8120-0
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DOI: https://doi.org/10.1007/s00253-017-8120-0