Increasing isobutanol yield by double-gene deletion of PDC6 and LPD1 in Saccharomyces cerevisiae

doi:10.1016/j.cjche.2016.04.004

›› 2016, Vol. 24 ›› Issue (8): 1074-1079.DOI: 10.1016/j.cjche.2016.04.004

• Biotechnology and Bioengineering • Previous Articles Next Articles

Increasing isobutanol yield by double-gene deletion of PDC6 and LPD1 in Saccharomyces cerevisiae

Aili Zhang, Yang Li, Yuhan Gao, Hongxing Jin

School of Chemical Engineering and Technology, Hebei University of Technology, Tianjin, 300130, China

Received:2015-06-18 Revised:2016-01-27 Online:2016-09-21 Published:2016-08-28
Supported by:
Supported by the National Natural Science Foundation of China (No. 21206028), the Doctoral Fund of Ministry of Education of China (No. 20121317120014), the Natural Science Foundation of Heibei Province (No. B2013202288), the Hebei Provincial Office of Education Science and Technology Research Projects (No. q2012024), the Hebei University of Technology Outstanding Youth Science and Technology Innovation Fund (No. 2012009) and the Open Fund of Key Laboratory of System Bioengineering of Ministry of Education of China (Tianjin University) (No. 20130315).

Increasing isobutanol yield by double-gene deletion of PDC6 and LPD1 in Saccharomyces cerevisiae

Aili Zhang, Yang Li, Yuhan Gao, Hongxing Jin

School of Chemical Engineering and Technology, Hebei University of Technology, Tianjin, 300130, China

通讯作者: Aili Zhang
基金资助:
Supported by the National Natural Science Foundation of China (No. 21206028), the Doctoral Fund of Ministry of Education of China (No. 20121317120014), the Natural Science Foundation of Heibei Province (No. B2013202288), the Hebei Provincial Office of Education Science and Technology Research Projects (No. q2012024), the Hebei University of Technology Outstanding Youth Science and Technology Innovation Fund (No. 2012009) and the Open Fund of Key Laboratory of System Bioengineering of Ministry of Education of China (Tianjin University) (No. 20130315).

Abstract

Abstract: As a new biofuel, isobutanol has received more attentions in recent years. Because of its high tolerance to higher alcohols, Saccharomyces cerevisiae has potential advantages as a platform microbe to produce isobutanol. In this study, we investigated integration effects of enhancing valine biosynthesis by overexpression of ILV2 and BAT2 with eliminating ethanol formation by deletion of PDC6 and decreasing acetyl-CoA biosynthesis by deletion of LPD1 on isobutanol titers. Our results showed that deletion of LPD1 in strains overexpressing BAT2 and ILV2 increased isobutanol titer by 5.3-fold compared with control strain. Additional deletion of PDC6 in lpd1Δ strains carrying overexpressed BAT2 and ILV2 further increased isobutanol titer by 1.5 fold. Overexpression of BAT2 and ILV2 in lpd1Δ strains and pdc6Δ strains decreased ethanol titers. Glycerol titers of the engineered strains did not have greater changes than that of control strain, while their acetic acid titers were higher, perhaps due to the imbalance of cofactors in isobutanol synthesis. Our researches suggest that double-gene deletion of PDC6 and LPD1 in strains overexpressing BAT2 and ILV2 could increase isobutanol production dramatically than single-gene deletion of PDC6 or LPD1. This study reveals the integration effects of overexpression of ILV2/BAT2 and double-gene deletion of LPD1 and PDC6 on isobutanol production, and helps understanding future developments of engineered strains for producing isobutanol.

Key words: Saccharomyces cerevisiae, Isobutanol, Ethanol, Pyruvate decarboxylase

摘要： As a new biofuel, isobutanol has received more attentions in recent years. Because of its high tolerance to higher alcohols, Saccharomyces cerevisiae has potential advantages as a platform microbe to produce isobutanol. In this study, we investigated integration effects of enhancing valine biosynthesis by overexpression of ILV2 and BAT2 with eliminating ethanol formation by deletion of PDC6 and decreasing acetyl-CoA biosynthesis by deletion of LPD1 on isobutanol titers. Our results showed that deletion of LPD1 in strains overexpressing BAT2 and ILV2 increased isobutanol titer by 5.3-fold compared with control strain. Additional deletion of PDC6 in lpd1Δ strains carrying overexpressed BAT2 and ILV2 further increased isobutanol titer by 1.5 fold. Overexpression of BAT2 and ILV2 in lpd1Δ strains and pdc6Δ strains decreased ethanol titers. Glycerol titers of the engineered strains did not have greater changes than that of control strain, while their acetic acid titers were higher, perhaps due to the imbalance of cofactors in isobutanol synthesis. Our researches suggest that double-gene deletion of PDC6 and LPD1 in strains overexpressing BAT2 and ILV2 could increase isobutanol production dramatically than single-gene deletion of PDC6 or LPD1. This study reveals the integration effects of overexpression of ILV2/BAT2 and double-gene deletion of LPD1 and PDC6 on isobutanol production, and helps understanding future developments of engineered strains for producing isobutanol.

关键词: Saccharomyces cerevisiae, Isobutanol, Ethanol, Pyruvate decarboxylase

Aili Zhang, Yang Li, Yuhan Gao, Hongxing Jin. Increasing isobutanol yield by double-gene deletion of PDC6 and LPD1 in Saccharomyces cerevisiae[J]. , 2016, 24(8): 1074-1079.

References

[1] K.Kuroda,M.Ueda,Cellular and molecular engineering of yeast Saccharomyces cerevisiae for advanced biobutanol production,FEMS Microbiol.Lett.363(3) (Feb 2016).
[2] C.Weber,A.Farwick,F.Benisch,et al.,Trends and challenges in the microbial production of lignocellulosic bioalcohol fuels,Appl.Microbiol.Biotechnol.87(4) (2010)1303-1315.
[3] H.Sakuragi,K.Kuroda,M.Ueda,Molecular breeding of advanced microorganisms for biofuel production,J.Biomed.Biotechnol.1(17)(2011)1-11.
[4] S.Atsumi,T.Hanai,J.C.Liao,Non-fermentative pathways for synthesis of branchedchain higher alcohols as biofuels,Nature 451(7174)(2008)86-89.
[5] B.Blombach,B.J.Eikmanns,Current knowledge on isobutanol production with Escherichia coli,Bacillus subtilis and Corynebacterium glutamicum,Bioeng.Bugs 2(6)(2011)346-350.
[6] W.Higashide,Y.Li,Y.Yang,J.C.Liao,Metabolic engineering of Clostridium cellulolyticum for production of isobutanol from cellulose,Appl.Environ.Microbiol. 77(8)(2011)2727-2733.
[7] S.Li,J.Wen,X.Jia,Engineering Bacillus subtilis for isobutanol production by heterologous Ehrlich pathway construction and the biosynthetic 2-ketoisovalerate precursor pathway overexpression,Appl.Microbiol.Biotechnol.91(3)(2011)577-589.
[8] K.M.Smith,K.M.Cho,J.C.Liao,Engineering Corynebacterium glutamicum for isobutanol production,Appl.Microbiol.Biotechnol.87(3)(2010)1045-1055.
[9] A.M.Varman,Y.Xiao,P.HB,et al.,Metabolic engineering of Synechocystis sp.strain PCC 6803 for isobutanol production,Appl.Environ.Microbiol.79(3)(2013)908-914.
[10] B.-R.Oh,S.-Y.Heo,S.-M.Lee,et al.,Erratum to production of isobutanol from crude glycerol by a genetically engineered Klebsiella pneumoniae strain,Biotechnol.Lett.10(25)(2013)1-6.
[11] M.X.He,B.Wu,H.Qin,et al.,Zymomonas mobilis:A novel platform for future biorefineries,Biotechnol.Biofuels 7(101)(2014)1-15.
[12] X.Chen,K.F.Nielsen,I.Borodina,et al.,Increased isobutanol production in Saccharomyces cerevisiae by overexpression of genes in valine metabolism,Biotechnol. Biofuels 4(21)(2011)1-12.
[13] K.K.Hong,Nielsen,Metabolic engineering of Saccharomyces cerevisiae:A key cell factory platform for future biorefineries,Cell.Mol.Life Sci.69(16)(2012) 2671-2690.
[14] E.P.Knoshaug,M.Zhang,Butanol tolerance in a selection of microorganisms,Appl. Biochem.Biotechnol.153(2009)13-20.
[15] P.Fatehi,Recent advancements in various steps of ethanol,butanol,and isobutanol productions from woody materials,Biotechnol.Prog.29(2)(2013)297-310.
[16] T. Kondo, H. Tezuka, J. Ishii, et al., Genetic engineering to enhance the Ehrlich pathway and alter carbon flux for increased isobutanol production from glucose by Saccharomyces cerevisiae, J. Biotechnol. 159(1-2) (2012) 32-37.
[17] W.-H. Lee, S.-O. Seo, Y.-H. Bae, et al., Isobutanol production in engineered Saccharomyces cerevisiae by overexpression of 2-ketoisovalerate decarboxylase and valine biosynthetic enzymes, Bioprocess Biosyst. Eng. 35(9) (2012) 1467-1475.
[18] E. Ofuonye, K. Kutin, D.T. Stuart, Engineering Saccharomyces cerevisiae fermentative pathways for the production of isobutanol, Biofuels 4(2) (2013) 185-201.
[19] D. Brat, E. Boles, Isobutanol production from D-xylose by recombinant Saccharomyces cerevisiae, FEMS Yeast Res. 13(2013) 241-244.
[20] D. Brat, C. Weber, W. Lorenzen, et al., Cytosolic re-localization and optimization of valine synthesis and catabolism enables increased isobutanol production with the yeast Saccharomyces cerevisiae, Biotechnol. Biofuels 5(65) (2012) 1-16.
[21] J.L. Avalos, G.R. Fink, G. Stephanopoulos, Compartmentalization of metabolic pathways in yeast mitochondria improves the production of branched-chain alcohols, Nat. Biotechnol. 31(2013) 335-341.
[22] K. Ida, J. Ishii, F. Matsuda, et al., Eliminating the isoleucine biosynthetic pathway to reduce competitive carbon outflow during isobutanol production by Saccharomyces cerevisiae, Microb. Cell Factories 14(2015) 62-70.
[23] F. Matsuda, J. Ishii, T. Kondo, et al., Increased isobutanol production in Saccharomy cescerevisiae by eliminating competing pathways and resolving cofactor imbalance, Microb. Cell Factories 12(2013) 119-129.
[24] B.J. Thomas, R. Rothstein, Elevated recombination rates in transcriptionally active DNA, Cell 56(4) (1989) 619-630.
[25] R.D. Gietz, A. Sugino, New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites, Gene 74(2) (1988) 527-534.
[26] A.L. Zhang, X. Chen, Improveethanol titer through minimizing glycerol titer in ethanol fermentation of Saccharomyces cerevisiae, Chin. J. Chem. Eng. 16(4) (2008) 620-625.

Increasing isobutanol yield by double-gene deletion of PDC6 and LPD1 in Saccharomyces cerevisiae

Increasing isobutanol yield by double-gene deletion of PDC6 and LPD1 in Saccharomyces cerevisiae

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	Tingjun Fu, Ran Wang, Kun Ren, Liangliang Zhang, Zhong Li. Intensified shape selectivity and alkylation reaction for the two-step conversion of methanol aromatization to p-xylene [J]. Chinese Journal of Chemical Engineering, 2023, 59(7): 240-250.
[2]	Yi Wu, Pengfei Song, Ningyan Li, Yanan Jiang, Yuan Liu. Molybdenum tailored Co⁰/Co²⁺ active pairs on a perovskite-type oxide for direct ethanol synthesis from syngas [J]. Chinese Journal of Chemical Engineering, 2023, 59(7): 279-289.
[3]	Pengcheng Zou, Kai Wang. Methanolysis of amides under high-temperature and high-pressure conditions with a continuous tubular reactor [J]. Chinese Journal of Chemical Engineering, 2023, 58(6): 170-178.
[4]	Jiaxin Wu, Chenxiao Wang, Xianliang Meng, Haichen Liu, Ruizhi Chu, Guoguang Wu, Weisong Li, Xiaofeng Jiang, Deguang Yang. Enhancement of catalytic and anti-carbon deposition performance of SAPO-34/ZSM-5/quartz films in MTA reaction by Si/Al ratio regulation [J]. Chinese Journal of Chemical Engineering, 2023, 56(4): 314-324.
[5]	Xinhai Zhou, Dawei Zhou, Xinhui Bao, Yang Zhang, Jie Zhou, Fengxue Xin, Wenming Zhang, Xiujuan Qian, Weiliang Dong, Min Jiang, Katrin Ochsenreither. Production of palmitoleic acid by oleaginous yeast Scheffersomyces segobiensis DSM 27193 using systematic dissolved oxygen regulation strategy [J]. Chinese Journal of Chemical Engineering, 2023, 53(1): 324-331.
[6]	Xing Su, Ning Qiao, Bao-Chang Sun. A route for the study on mass transfer enhancement by adding particles in liquid phase [J]. Chinese Journal of Chemical Engineering, 2022, 48(8): 158-165.
[7]	Siyue Ren, Xiao Feng. Emergy evaluation of aromatics production from methanol and naphtha [J]. Chinese Journal of Chemical Engineering, 2022, 46(6): 134-141.
[8]	Xiangjun Li, Shujun Li, Xiaoping Wang, Muhammad Asif Nawaz, Dianhua Liu. Polyoxymethylene dimethyl ethers synthesis from methanol and formaldehyde solution over one-pot synthesized spherical mesoporous sulfated zirconia [J]. Chinese Journal of Chemical Engineering, 2022, 46(6): 161-172.
[9]	Lijuan He, Cuimei Zhi, Lixia Ling, Riguang Zhang, Baojun Wang. Syngas to ethanol on MoCu(2 1 1) surface: Effect of promoter Mo on C—O bond breaking and C—C bond formation [J]. Chinese Journal of Chemical Engineering, 2022, 45(5): 78-89.
[10]	Youwei Yang, Jingyu Zhang, Yueqi Gao, Busha Assaba Fayisa, Antai Li, Shouying Huang, Jing Lv, Yue Wang, Xinbin Ma. Highly dispersed nickel boosts catalysis by Cu/SiO₂ in the hydrogenation of CO₂-derived ethylene carbonate to methanol and ethylene glycol [J]. Chinese Journal of Chemical Engineering, 2022, 43(3): 77-85.
[11]	Yihan Yin, Aoqian Qiu, Hongxia Gao, Yanqing Na, Zhiwu Liang. Experimental study of the mass transfer behavior of carbon dioxide absorption into ternary phase change solution in a packed tower [J]. Chinese Journal of Chemical Engineering, 2022, 43(3): 135-142.
[12]	Wanqiao Liang, Jihuan Huang, Penny Xiao, Ranjeet Singh, Jining Guo, Leila Dehdari, Gang Kevin Li. Amine-immobilized HY zeolite for CO₂ capture from hot flue gas [J]. Chinese Journal of Chemical Engineering, 2022, 43(3): 335-342.
[13]	Xia Zhan, Xueying Zhao, Zhongyong Gao, Rui Ge, Juan Lu, Luying Wang, Jiding Li. Breakthroughs on tailoring membrane materials for ethanol recovery by pervaporation [J]. Chinese Journal of Chemical Engineering, 2022, 52(12): 19-36.
[14]	Busha Assaba Fayisa, Yushan Xi, Youwei Yang, Yueqi Gao, Antai Li, Mei-Yan Wang, Jing Lv, Shouying Huang, Yue Wang, Xinbin Ma. Pt-modulated Cu/SiO₂ catalysts for efficient hydrogenation of CO₂-derived ethylene carbonate to methanol and ethylene glycol [J]. Chinese Journal of Chemical Engineering, 2022, 41(1): 366-373.
[15]	Yu Dong, Tiantian Ping, Shufeng Shen. Solubility of CO₂ in nonaqueous system of 2-(butylamino)ethanol with 2-butoxyethanol: Experimental data and model representation [J]. Chinese Journal of Chemical Engineering, 2022, 41(1): 441-448.