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Shangtian Yang - One of the best experts on this subject based on the ideXlab platform.
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Metabolic engineering strategies for Acetoin and 2,3-butanediol production: advances and prospects.
Critical reviews in biotechnology, 2017Co-Authors: Taowei Yang, Xian Zhang, Zhiming Rao, Shangtian YangAbstract:Acetoin and 2,3-butanediol (2,3-BD) have a large number of industrial applications. The production of Acetoin and 2,3-BD has traditionally relied on oil supplies. Microbial production of Acetoin and 2,3-BD will alleviate the dependence on oil. Acetoin and 2,3-BD are neighboring metabolites in the 2,3-BD metabolic pathway of bacteria. This review summarizes metabolic engineering strategies for improvement of microbial Acetoin and 2,3-BD production. We also propose enhancements to current Acetoin and 2,3-BD production strategies, by offering a metabolic engineering approach that is guided by systems biology and synthetic biology.
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efficient whole cell biocatalyst for Acetoin production with nad regeneration system through homologous co expression of 2 3 butanediol dehydrogenase and nadh oxidase in engineered bacillus subtilis
PLOS ONE, 2014Co-Authors: Xian Zhang, Xiaojing Zhao, Rongzhen Zhang, Taowei Yang, Zhenghong Xu, Shangtian YangAbstract:Acetoin (3-hydroxy-2-butanone), an extensively-used food spice and bio-based platform chemical, is usually produced by chemical synthesis methods. With increasingly requirement of food security and environmental protection, bio-fermentation of Acetoin by microorganisms has a great promising market. However, through metabolic engineering strategies, the mixed acid-butanediol fermentation metabolizes a certain portion of substrate to the by-products of organic acids such as lactic acid and acetic acid, which causes energy cost and increases the difficulty of product purification in downstream processes. In this work, due to the high efficiency of enzymatic reaction and excellent selectivity, a strategy for efficiently converting 2,3-butandiol to Acetoin using whole-cell biocatalyst by engineered Bacillus subtilis is proposed. In this process, NAD+ plays a significant role on 2,3-butanediol and Acetoin distribution, so the NADH oxidase and 2,3-butanediol dehydrogenase both from B. subtilis are co-expressed in B. subtilis 168 to construct an NAD+ regeneration system, which forces dramatic decrease of the intracellular NADH concentration (1.6 fold) and NADH/NAD+ ratio (2.2 fold). By optimization of the enzymatic reaction and applying repeated batch conversion, the whole-cell biocatalyst efficiently produced 91.8 g/L Acetoin with a productivity of 2.30 g/(L·h), which was the highest record ever reported by biocatalysis. This work indicated that manipulation of the intracellular cofactor levels was more effective than the strategy of enhancing enzyme activity, and the bioprocess for NAD+ regeneration may also be a useful way for improving the productivity of NAD+-dependent chemistry-based products.
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the rebalanced pathway significantly enhances Acetoin production by disruption of Acetoin reductase gene and moderate expression of a new water forming nadh oxidase in bacillus subtilis
Metabolic Engineering, 2014Co-Authors: Xian Zhang, Rongzhen Zhang, Taowei Yang, Zhiming Rao, Teng Bao, Shangtian YangAbstract:Bacillus subtilis produces Acetoin as a major extracellular product. However, the by-products of 2,3-butanediol, lactic acid and ethanol were accompanied in the NADH-dependent pathways. In this work, metabolic engineering strategies were proposed to redistribute the carbon flux to Acetoin by manipulation the NADH levels. We first knocked out the Acetoin reductase gene bdhA to block the main flux from Acetoin to 2,3-butanediol. Then, among four putative candidates, we successfully screened an active water-forming NADH oxidase, YODC. Moderate-expression of YODC in the bdhA disrupted B. subtilis weakened the NADH-linked pathways to by-product pools of Acetoin. Through these strategies, Acetoin production was improved to 56.7g/l with an increase of 35.3%, while the production of 2,3-butanediol, lactic acid and ethanol were decreased by 92.3%, 70.1% and 75.0%, respectively, simultaneously the fermentation duration was decreased 1.7-fold. Acetoin productivity by B. subtilis was improved to 0.639g/(lh).
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two stage ph control strategy based on the ph preference of Acetoin reductase regulates Acetoin and 2 3 butanediol distribution in bacillus subtilis
PLOS ONE, 2014Co-Authors: Xian Zhang, Taowei Yang, Zhenghong Xu, Shangtian Yang, Huazhong LiAbstract:Acetoin reductase/2,3-butanediol dehydrogenase (AR/BDH), which catalyzes the interconversion between Acetoin and 2,3-butanediol, plays an important role in distribution of the products pools. This work characterized the Bacillus subtilis AR/BDH for the first time. The enzyme showed very different pH preferences of pH 6.5 for reduction and pH 8.5 for oxidation. Based on these above results, a two-stage pH control strategy was optimized for Acetoin production, in which the pH was controlled at 6.5 for quickly converting glucose to Acetoin and 2,3-butanediol, and then 8.0 for reversely transforming 2,3-butanediol to Acetoin. By over-expression of AR/BDH in the wild-type B. subtilis JNA 3-10 and applying fed-batch fermentation based on the two-stage pH control strategy, Acetoin yield of B. subtilis was improved to a new record of 73.6 g/l, with the productivity of 0.77 g/(l·h). The molar yield of Acetoin was improved from 57.5% to 83.5% and the ratio of Acetoin/2,3-butanediol was switched from 2.7∶1 to 18.0∶1.
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effects of corn steep liquor on production of 2 3 butanediol and Acetoin by bacillus subtilis
Process Biochemistry, 2013Co-Authors: Taowei Yang, Xian Zhang, Zhenghong Xu, Meijuan Xu, Shangtian YangAbstract:Abstract The initial concentration of corn steep liquor (CSL) have remarkable effects on not only 2,3-butanediol (2,3-BD) and Acetoin (metabolic precursor) production, but also on the ratio of 2,3-BD to Acetoin. When a high concentration of CSL was supplemented, cell growth was improved, Acetoin reductase (ACR) was stimulated, the concentration of 2,3-BD increased by 78.6%, Acetoin decreased by 61.9%, and the ratio of 2,3-BD to Acetoin increased by 3.69-fold. The acr gene, encoding ACR, was over-expressed in Bacillus subtilis . Compared to the control (parent strain), low levels of CSL in the engineered strain increased 2,3-BD concentration and the ratio 2,3-BD to Acetoin by 13.9% and 39.5%, respectively, and decreased Acetoin titer by 18.3%. Acetoin became a major product under low levels of CSL. Also, a knockout strain carrying an acr::cat insertion mutation was constructed. As expected, the loss of ACR activity led to an accumulation of Acetoin in the supernatants of acr:: cat mutant cultures. Additionally, the productivity of Acetoin was improved by high concentration of CSL. The results above demonstrate the feasibility of using B. subtilis for the production of not only 2,3-BD but also Acetoin as a major product.
Liaoyuan Zhang - One of the best experts on this subject based on the ideXlab platform.
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an artificial synthetic pathway for Acetoin 2 3 butanediol and 2 butanol production from ethanol using cell free multi enzyme catalysis
Green Chemistry, 2018Co-Authors: Liaoyuan Zhang, Raushan Kumar Singh, Dakshinamurthy Sivakumar, Zewang Guo, Fanbing Chen, Xiong Guan, Yun Chan Kang, Jungkul LeeAbstract:Upgrading ethanol to higher order alcohols is desired but difficult using current biotechnological methods. In this study, we designed a completely artificial reaction pathway for upgrading ethanol to Acetoin, 2,3-butanediol, and 2-butanol in a cell-free bio-system composed of ethanol dehydrogenase, formolase, 2,3-butanediol dehydrogenase, diol dehydratase, and NADH oxidase. Under optimized conditions, Acetoin, 2,3-butanediol, and 2-butanol were produced at 88.78%, 88.28%, and 27.25% of the theoretical yield from 100 mM ethanol, respectively. These results demonstrate that this artificial synthetic pathway is an environmentally-friendly novel approach for upgrading bio-ethanol to Acetoin, 2,3-butanediol, and 2-butanol.
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Efficient (3S)-Acetoin and (2S,3S)-2,3-Butanediol Production from meso-2,3-Butanediol Using Whole-Cell Biocatalysis
MDPI AG, 2018Co-Authors: Feixue Chen, Zewang Guo, Meijing Sun, Huifang Gao, Hui Lin, Jiebo Chen, Wensong Jin, Yunlong Yang, Liaoyuan ZhangAbstract:(3S)-Acetoin and (2S,3S)-2,3-butanediol are important platform chemicals widely applied in the asymmetric synthesis of valuable chiral chemicals. However, their production by fermentative methods is difficult to perform. This study aimed to develop a whole-cell biocatalysis strategy for the production of (3S)-Acetoin and (2S,3S)-2,3-butanediol from meso-2,3-butanediol. First, E. coli co-expressing (2R,3R)-2,3-butanediol dehydrogenase, NADH oxidase and Vitreoscilla hemoglobin was developed for (3S)-Acetoin production from meso-2,3-butanediol. Maximum (3S)-Acetoin concentration of 72.38 g/L with the stereoisomeric purity of 94.65% was achieved at 24 h under optimal conditions. Subsequently, we developed another biocatalyst co-expressing (2S,3S)-2,3-butanediol dehydrogenase and formate dehydrogenase for (2S,3S)-2,3-butanediol production from (3S)-Acetoin. Synchronous catalysis together with two biocatalysts afforded 38.41 g/L of (2S,3S)-butanediol with stereoisomeric purity of 98.03% from 40 g/L meso-2,3-butanediol. These results exhibited the potential for (3S)-Acetoin and (2S,3S)-butanediol production from meso-2,3-butanediol as a substrate via whole-cell biocatalysis
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a new nad h dependent meso 2 3 butanediol dehydrogenase from an industrially potential strain serratia marcescens h30
Applied Microbiology and Biotechnology, 2014Co-Authors: Liaoyuan Zhang, Xiong Guan, Quanming Xu, Senran Zhan, Yongyu Li, Kaihui Hu, Yaling ShenAbstract:The budC gene coding for a new meso-2,3-butanediol dehydrogenase (BDH) from Serratia marcescens H30 was cloned and expressed in Escherichia coli BL21(DE3), purified, and characterized for its properties. The recombinant BDH with a molecular weight of 27.4 kDa exhibited a reversible transformation between Acetoin and 2,3-butanediol. In the presence of NADH, BDH could catalyze the reduction of diacetyl and (3R)-Acetoin to (3S)-Acetoin and meso-2,3-butanediol, respectively, while (3S)-Acetoin as a substrate could be further transformed into (2S, 3S)-2,3-butanediol at pH 9.0. For diol oxidation reactions, (3R)-Acetoin and (3S)-Acetoin were obtained when meso-2,3-butanediol and (2S,3S)-2,3-butanediol were used as the substrates with BDH and NAD+. (2R,3R)-2,3-butanediol was not a substrate for the BDH at all. The low Km value (4.1 mM) in meso-2,3-butanediol oxidation reaction and no activity for diacetyl, Acetoin, and 2,3-butanediol as the substrates with NADP+/NADPH suggested that the budC gene product belongs to a NAD(H)-dependent meso-2,3-BDH. Maximum activities for diacetyl and (3S/3R)-Acetoin reduction were observed at pH 8.0 and pH 5.0 while for meso-2,3-butanediol oxidation it was pH 8.0. However, the optimum temperature for oxidation and reduction reactions was about 40 °C. In addition, the BDH activity for meso-2,3-butanediol oxidation was enhanced in the presence of Fe2+ and for diacetyl and (3S/3R)-Acetoin reduction in the presence of Mg2+ and Mn2+, while several metal ions inhibited its activity, particularly Fe3+ for reduction of diacetyl and Acetoin. Sequence analysis showed that the BDH from S. marcescens H30 possessed two conserved sequences including the coenzyme binding motif (GxxxGxG) and the active-site motif (YxxxK), which are present in the short-chain dehydrogenase/reductase superfamily.
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efficient Acetoin production by optimization of medium components and oxygen supply control using a newly isolated paenibacillus polymyxa cs107
Journal of Chemical Technology & Biotechnology, 2012Co-Authors: Liaoyuan Zhang, Shuang Chen, Haibo Xie, Yuting TianAbstract:BACKGROUND: Acetoin is a natural flavor commonly used in wine, buffer, honey, garnet berry and strawberry as a food additive. It also has been widely applied in cosmetics, pharmacy and chemical synthesis. Culture medium optimization and process control were carried out for efficient production of Acetoin by a newly isoliated P. polymyxa CS107. RESULTS: An Acetoin high producing strain, designated as CS107, was newly isolated and identified as P. polymyxa based on its physiological and biochemical characteristics as well as the 16S rDNA sequence. The medium composition was optimized in shake flask fermentations by a sequential statistical experimental design. Under the optimized conditions, Acetoin concentration of 30.98 g L−1 was achieved with 71.83% of theoretical glucose conversion efficiency. Fed-batch fermentation based on a suitable agitation speed was carried out in a 5 L jar, the maximum Acetoin concentration of 55.3 g L−1 was obtained with the productivity of 1.32 g L−1 h−1 and the yield of 75.62%. CONCLUSION: A new strain for efficient production of Acetoin, designated as P. polymyxa CS107, was obtained. The optimization of fermentation variables and fed-batch culture resulted in a maximum Acetoin concentration of 55.3 g L−1 in 5 L jar. Copyright © 2012 Society of Chemical Industry
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enhanced Acetoin production by serratia marcescens h32 with expression of a water forming nadh oxidase
Bioresource Technology, 2012Co-Authors: Liaoyuan Zhang, Yaling ShenAbstract:Abstract Cofactor engineering was employed to enhance production of Acetoin by Serratia marcescens H32. 2,3-Butanediol was a major byproduct of Acetoin fermentation by S. marcescens H32. In order to decrease 2,3-butanediol formation and achieve a high efficiency of Acetoin production, nox gene encoding a water-forming NADH oxidase from Lactobacillus brevis was expressed. Batch fermentations suggested the expression of the NADH oxidase could increase the intracellular NAD + concentration (1.5-fold) and NAD + /NADH ratio (2.9-fold). Meanwhile, 2,3-butanediol was significantly decreased (52%), and the accumulation of Acetoin was enhanced (33%) accordingly. By fed-batch culture of the engineered strain, the final Acetoin titer up to 75.2 g/l with the productivity of 1.88 g/(l h) was obtained. To the best of our knowledge, these results were new records on Acetoin fermentation ever reported.
Taowei Yang - One of the best experts on this subject based on the ideXlab platform.
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Metabolic engineering strategies for Acetoin and 2,3-butanediol production: advances and prospects.
Critical reviews in biotechnology, 2017Co-Authors: Taowei Yang, Xian Zhang, Zhiming Rao, Shangtian YangAbstract:Acetoin and 2,3-butanediol (2,3-BD) have a large number of industrial applications. The production of Acetoin and 2,3-BD has traditionally relied on oil supplies. Microbial production of Acetoin and 2,3-BD will alleviate the dependence on oil. Acetoin and 2,3-BD are neighboring metabolites in the 2,3-BD metabolic pathway of bacteria. This review summarizes metabolic engineering strategies for improvement of microbial Acetoin and 2,3-BD production. We also propose enhancements to current Acetoin and 2,3-BD production strategies, by offering a metabolic engineering approach that is guided by systems biology and synthetic biology.
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efficient whole cell biocatalyst for Acetoin production with nad regeneration system through homologous co expression of 2 3 butanediol dehydrogenase and nadh oxidase in engineered bacillus subtilis
PLOS ONE, 2014Co-Authors: Xian Zhang, Xiaojing Zhao, Rongzhen Zhang, Taowei Yang, Zhenghong Xu, Shangtian YangAbstract:Acetoin (3-hydroxy-2-butanone), an extensively-used food spice and bio-based platform chemical, is usually produced by chemical synthesis methods. With increasingly requirement of food security and environmental protection, bio-fermentation of Acetoin by microorganisms has a great promising market. However, through metabolic engineering strategies, the mixed acid-butanediol fermentation metabolizes a certain portion of substrate to the by-products of organic acids such as lactic acid and acetic acid, which causes energy cost and increases the difficulty of product purification in downstream processes. In this work, due to the high efficiency of enzymatic reaction and excellent selectivity, a strategy for efficiently converting 2,3-butandiol to Acetoin using whole-cell biocatalyst by engineered Bacillus subtilis is proposed. In this process, NAD+ plays a significant role on 2,3-butanediol and Acetoin distribution, so the NADH oxidase and 2,3-butanediol dehydrogenase both from B. subtilis are co-expressed in B. subtilis 168 to construct an NAD+ regeneration system, which forces dramatic decrease of the intracellular NADH concentration (1.6 fold) and NADH/NAD+ ratio (2.2 fold). By optimization of the enzymatic reaction and applying repeated batch conversion, the whole-cell biocatalyst efficiently produced 91.8 g/L Acetoin with a productivity of 2.30 g/(L·h), which was the highest record ever reported by biocatalysis. This work indicated that manipulation of the intracellular cofactor levels was more effective than the strategy of enhancing enzyme activity, and the bioprocess for NAD+ regeneration may also be a useful way for improving the productivity of NAD+-dependent chemistry-based products.
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the rebalanced pathway significantly enhances Acetoin production by disruption of Acetoin reductase gene and moderate expression of a new water forming nadh oxidase in bacillus subtilis
Metabolic Engineering, 2014Co-Authors: Xian Zhang, Rongzhen Zhang, Taowei Yang, Zhiming Rao, Teng Bao, Shangtian YangAbstract:Bacillus subtilis produces Acetoin as a major extracellular product. However, the by-products of 2,3-butanediol, lactic acid and ethanol were accompanied in the NADH-dependent pathways. In this work, metabolic engineering strategies were proposed to redistribute the carbon flux to Acetoin by manipulation the NADH levels. We first knocked out the Acetoin reductase gene bdhA to block the main flux from Acetoin to 2,3-butanediol. Then, among four putative candidates, we successfully screened an active water-forming NADH oxidase, YODC. Moderate-expression of YODC in the bdhA disrupted B. subtilis weakened the NADH-linked pathways to by-product pools of Acetoin. Through these strategies, Acetoin production was improved to 56.7g/l with an increase of 35.3%, while the production of 2,3-butanediol, lactic acid and ethanol were decreased by 92.3%, 70.1% and 75.0%, respectively, simultaneously the fermentation duration was decreased 1.7-fold. Acetoin productivity by B. subtilis was improved to 0.639g/(lh).
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two stage ph control strategy based on the ph preference of Acetoin reductase regulates Acetoin and 2 3 butanediol distribution in bacillus subtilis
PLOS ONE, 2014Co-Authors: Xian Zhang, Taowei Yang, Zhenghong Xu, Shangtian Yang, Huazhong LiAbstract:Acetoin reductase/2,3-butanediol dehydrogenase (AR/BDH), which catalyzes the interconversion between Acetoin and 2,3-butanediol, plays an important role in distribution of the products pools. This work characterized the Bacillus subtilis AR/BDH for the first time. The enzyme showed very different pH preferences of pH 6.5 for reduction and pH 8.5 for oxidation. Based on these above results, a two-stage pH control strategy was optimized for Acetoin production, in which the pH was controlled at 6.5 for quickly converting glucose to Acetoin and 2,3-butanediol, and then 8.0 for reversely transforming 2,3-butanediol to Acetoin. By over-expression of AR/BDH in the wild-type B. subtilis JNA 3-10 and applying fed-batch fermentation based on the two-stage pH control strategy, Acetoin yield of B. subtilis was improved to a new record of 73.6 g/l, with the productivity of 0.77 g/(l·h). The molar yield of Acetoin was improved from 57.5% to 83.5% and the ratio of Acetoin/2,3-butanediol was switched from 2.7∶1 to 18.0∶1.
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effects of corn steep liquor on production of 2 3 butanediol and Acetoin by bacillus subtilis
Process Biochemistry, 2013Co-Authors: Taowei Yang, Xian Zhang, Zhenghong Xu, Meijuan Xu, Shangtian YangAbstract:Abstract The initial concentration of corn steep liquor (CSL) have remarkable effects on not only 2,3-butanediol (2,3-BD) and Acetoin (metabolic precursor) production, but also on the ratio of 2,3-BD to Acetoin. When a high concentration of CSL was supplemented, cell growth was improved, Acetoin reductase (ACR) was stimulated, the concentration of 2,3-BD increased by 78.6%, Acetoin decreased by 61.9%, and the ratio of 2,3-BD to Acetoin increased by 3.69-fold. The acr gene, encoding ACR, was over-expressed in Bacillus subtilis . Compared to the control (parent strain), low levels of CSL in the engineered strain increased 2,3-BD concentration and the ratio 2,3-BD to Acetoin by 13.9% and 39.5%, respectively, and decreased Acetoin titer by 18.3%. Acetoin became a major product under low levels of CSL. Also, a knockout strain carrying an acr::cat insertion mutation was constructed. As expected, the loss of ACR activity led to an accumulation of Acetoin in the supernatants of acr:: cat mutant cultures. Additionally, the productivity of Acetoin was improved by high concentration of CSL. The results above demonstrate the feasibility of using B. subtilis for the production of not only 2,3-BD but also Acetoin as a major product.
Xian Zhang - One of the best experts on this subject based on the ideXlab platform.
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Metabolic engineering strategies for Acetoin and 2,3-butanediol production: advances and prospects.
Critical reviews in biotechnology, 2017Co-Authors: Taowei Yang, Xian Zhang, Zhiming Rao, Shangtian YangAbstract:Acetoin and 2,3-butanediol (2,3-BD) have a large number of industrial applications. The production of Acetoin and 2,3-BD has traditionally relied on oil supplies. Microbial production of Acetoin and 2,3-BD will alleviate the dependence on oil. Acetoin and 2,3-BD are neighboring metabolites in the 2,3-BD metabolic pathway of bacteria. This review summarizes metabolic engineering strategies for improvement of microbial Acetoin and 2,3-BD production. We also propose enhancements to current Acetoin and 2,3-BD production strategies, by offering a metabolic engineering approach that is guided by systems biology and synthetic biology.
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efficient whole cell biocatalyst for Acetoin production with nad regeneration system through homologous co expression of 2 3 butanediol dehydrogenase and nadh oxidase in engineered bacillus subtilis
PLOS ONE, 2014Co-Authors: Xian Zhang, Xiaojing Zhao, Rongzhen Zhang, Taowei Yang, Zhenghong Xu, Shangtian YangAbstract:Acetoin (3-hydroxy-2-butanone), an extensively-used food spice and bio-based platform chemical, is usually produced by chemical synthesis methods. With increasingly requirement of food security and environmental protection, bio-fermentation of Acetoin by microorganisms has a great promising market. However, through metabolic engineering strategies, the mixed acid-butanediol fermentation metabolizes a certain portion of substrate to the by-products of organic acids such as lactic acid and acetic acid, which causes energy cost and increases the difficulty of product purification in downstream processes. In this work, due to the high efficiency of enzymatic reaction and excellent selectivity, a strategy for efficiently converting 2,3-butandiol to Acetoin using whole-cell biocatalyst by engineered Bacillus subtilis is proposed. In this process, NAD+ plays a significant role on 2,3-butanediol and Acetoin distribution, so the NADH oxidase and 2,3-butanediol dehydrogenase both from B. subtilis are co-expressed in B. subtilis 168 to construct an NAD+ regeneration system, which forces dramatic decrease of the intracellular NADH concentration (1.6 fold) and NADH/NAD+ ratio (2.2 fold). By optimization of the enzymatic reaction and applying repeated batch conversion, the whole-cell biocatalyst efficiently produced 91.8 g/L Acetoin with a productivity of 2.30 g/(L·h), which was the highest record ever reported by biocatalysis. This work indicated that manipulation of the intracellular cofactor levels was more effective than the strategy of enhancing enzyme activity, and the bioprocess for NAD+ regeneration may also be a useful way for improving the productivity of NAD+-dependent chemistry-based products.
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the rebalanced pathway significantly enhances Acetoin production by disruption of Acetoin reductase gene and moderate expression of a new water forming nadh oxidase in bacillus subtilis
Metabolic Engineering, 2014Co-Authors: Xian Zhang, Rongzhen Zhang, Taowei Yang, Zhiming Rao, Teng Bao, Shangtian YangAbstract:Bacillus subtilis produces Acetoin as a major extracellular product. However, the by-products of 2,3-butanediol, lactic acid and ethanol were accompanied in the NADH-dependent pathways. In this work, metabolic engineering strategies were proposed to redistribute the carbon flux to Acetoin by manipulation the NADH levels. We first knocked out the Acetoin reductase gene bdhA to block the main flux from Acetoin to 2,3-butanediol. Then, among four putative candidates, we successfully screened an active water-forming NADH oxidase, YODC. Moderate-expression of YODC in the bdhA disrupted B. subtilis weakened the NADH-linked pathways to by-product pools of Acetoin. Through these strategies, Acetoin production was improved to 56.7g/l with an increase of 35.3%, while the production of 2,3-butanediol, lactic acid and ethanol were decreased by 92.3%, 70.1% and 75.0%, respectively, simultaneously the fermentation duration was decreased 1.7-fold. Acetoin productivity by B. subtilis was improved to 0.639g/(lh).
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two stage ph control strategy based on the ph preference of Acetoin reductase regulates Acetoin and 2 3 butanediol distribution in bacillus subtilis
PLOS ONE, 2014Co-Authors: Xian Zhang, Taowei Yang, Zhenghong Xu, Shangtian Yang, Huazhong LiAbstract:Acetoin reductase/2,3-butanediol dehydrogenase (AR/BDH), which catalyzes the interconversion between Acetoin and 2,3-butanediol, plays an important role in distribution of the products pools. This work characterized the Bacillus subtilis AR/BDH for the first time. The enzyme showed very different pH preferences of pH 6.5 for reduction and pH 8.5 for oxidation. Based on these above results, a two-stage pH control strategy was optimized for Acetoin production, in which the pH was controlled at 6.5 for quickly converting glucose to Acetoin and 2,3-butanediol, and then 8.0 for reversely transforming 2,3-butanediol to Acetoin. By over-expression of AR/BDH in the wild-type B. subtilis JNA 3-10 and applying fed-batch fermentation based on the two-stage pH control strategy, Acetoin yield of B. subtilis was improved to a new record of 73.6 g/l, with the productivity of 0.77 g/(l·h). The molar yield of Acetoin was improved from 57.5% to 83.5% and the ratio of Acetoin/2,3-butanediol was switched from 2.7∶1 to 18.0∶1.
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effects of corn steep liquor on production of 2 3 butanediol and Acetoin by bacillus subtilis
Process Biochemistry, 2013Co-Authors: Taowei Yang, Xian Zhang, Zhenghong Xu, Meijuan Xu, Shangtian YangAbstract:Abstract The initial concentration of corn steep liquor (CSL) have remarkable effects on not only 2,3-butanediol (2,3-BD) and Acetoin (metabolic precursor) production, but also on the ratio of 2,3-BD to Acetoin. When a high concentration of CSL was supplemented, cell growth was improved, Acetoin reductase (ACR) was stimulated, the concentration of 2,3-BD increased by 78.6%, Acetoin decreased by 61.9%, and the ratio of 2,3-BD to Acetoin increased by 3.69-fold. The acr gene, encoding ACR, was over-expressed in Bacillus subtilis . Compared to the control (parent strain), low levels of CSL in the engineered strain increased 2,3-BD concentration and the ratio 2,3-BD to Acetoin by 13.9% and 39.5%, respectively, and decreased Acetoin titer by 18.3%. Acetoin became a major product under low levels of CSL. Also, a knockout strain carrying an acr::cat insertion mutation was constructed. As expected, the loss of ACR activity led to an accumulation of Acetoin in the supernatants of acr:: cat mutant cultures. Additionally, the productivity of Acetoin was improved by high concentration of CSL. The results above demonstrate the feasibility of using B. subtilis for the production of not only 2,3-BD but also Acetoin as a major product.
Yaling Shen - One of the best experts on this subject based on the ideXlab platform.
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a new nad h dependent meso 2 3 butanediol dehydrogenase from an industrially potential strain serratia marcescens h30
Applied Microbiology and Biotechnology, 2014Co-Authors: Liaoyuan Zhang, Xiong Guan, Quanming Xu, Senran Zhan, Yongyu Li, Kaihui Hu, Yaling ShenAbstract:The budC gene coding for a new meso-2,3-butanediol dehydrogenase (BDH) from Serratia marcescens H30 was cloned and expressed in Escherichia coli BL21(DE3), purified, and characterized for its properties. The recombinant BDH with a molecular weight of 27.4 kDa exhibited a reversible transformation between Acetoin and 2,3-butanediol. In the presence of NADH, BDH could catalyze the reduction of diacetyl and (3R)-Acetoin to (3S)-Acetoin and meso-2,3-butanediol, respectively, while (3S)-Acetoin as a substrate could be further transformed into (2S, 3S)-2,3-butanediol at pH 9.0. For diol oxidation reactions, (3R)-Acetoin and (3S)-Acetoin were obtained when meso-2,3-butanediol and (2S,3S)-2,3-butanediol were used as the substrates with BDH and NAD+. (2R,3R)-2,3-butanediol was not a substrate for the BDH at all. The low Km value (4.1 mM) in meso-2,3-butanediol oxidation reaction and no activity for diacetyl, Acetoin, and 2,3-butanediol as the substrates with NADP+/NADPH suggested that the budC gene product belongs to a NAD(H)-dependent meso-2,3-BDH. Maximum activities for diacetyl and (3S/3R)-Acetoin reduction were observed at pH 8.0 and pH 5.0 while for meso-2,3-butanediol oxidation it was pH 8.0. However, the optimum temperature for oxidation and reduction reactions was about 40 °C. In addition, the BDH activity for meso-2,3-butanediol oxidation was enhanced in the presence of Fe2+ and for diacetyl and (3S/3R)-Acetoin reduction in the presence of Mg2+ and Mn2+, while several metal ions inhibited its activity, particularly Fe3+ for reduction of diacetyl and Acetoin. Sequence analysis showed that the BDH from S. marcescens H30 possessed two conserved sequences including the coenzyme binding motif (GxxxGxG) and the active-site motif (YxxxK), which are present in the short-chain dehydrogenase/reductase superfamily.
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enhanced Acetoin production by serratia marcescens h32 with expression of a water forming nadh oxidase
Bioresource Technology, 2012Co-Authors: Liaoyuan Zhang, Yaling ShenAbstract:Abstract Cofactor engineering was employed to enhance production of Acetoin by Serratia marcescens H32. 2,3-Butanediol was a major byproduct of Acetoin fermentation by S. marcescens H32. In order to decrease 2,3-butanediol formation and achieve a high efficiency of Acetoin production, nox gene encoding a water-forming NADH oxidase from Lactobacillus brevis was expressed. Batch fermentations suggested the expression of the NADH oxidase could increase the intracellular NAD + concentration (1.5-fold) and NAD + /NADH ratio (2.9-fold). Meanwhile, 2,3-butanediol was significantly decreased (52%), and the accumulation of Acetoin was enhanced (33%) accordingly. By fed-batch culture of the engineered strain, the final Acetoin titer up to 75.2 g/l with the productivity of 1.88 g/(l h) was obtained. To the best of our knowledge, these results were new records on Acetoin fermentation ever reported.
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enhanced Acetoin production by serratia marcescens h32 using statistical optimization and a two stage agitation speed control strategy
Biotechnology and Bioprocess Engineering, 2012Co-Authors: Liaoyuan Zhang, Yaling ShenAbstract:Enhanced Acetoin production was carried out by Serratia marcescens H32. First, medium compositions were optimized statistically for shake flask fermentations to produce Acetoin. Sucrose and corn steep liquor powder (CSLP) were identified as the most significant factors by Plackett-Burman design. The path of steepest ascent and response surface methodology were then employed to determine the optimal concentrations of the two factors. Acetoin yield was up to 41.5 g/L in flask fermentations using the optimized medium. Furthermore, the optimal medium was used to conduct fermentation experiments in a 3.7-L bioreactor. The influences of different agitation speeds on Acetoin production were investigated. Based on a process analysis, a two-stage agitation speed control strategy was proposed, in which the agitation speed was controlled at 700 rpm during the first 8 h and then switched to 600 rpm. A relatively high Acetoin concentration (44.9 g/L) and high Acetoin productivity (1.73 g/L/h) were achieved by applying this strategy. Fed-batch fermentation based on the two-stage agitation speed control strategy was performed, and a maximum Acetoin concentration of 60.5 g/L with productivity of 1.44 g/L/h was achieved.
