Abstract
Ionic liquids (ILs) are benign solvents that are highly effective for biomass pretreatment. However, their applications for scale-up biorefinery are limited due to multiple expensive IL recovery and separation steps that are required. To overcome this limitation, it is very critical to develop a compatible enzymatic and microbial biocatalyst system to carry the simultaneous saccharification and fermentation in IL environments (SSF-IL). While enzymatic biocatalysts have been demonstrated to be compatible with various IL environments, it is challenging to develop microbial biocatalysts that can thrive and perform efficient biotransformation under the same conditions (pH and temperature). In this study, we harnessed the robust metabolism of Yarrowia lipolytica as a microbial platform highly compatible with the IL environments such as 1-ethyl-3-methylimidazolium acetate ([EMIM][OAc]). We optimized the enzymatic and microbial biocatalyst system using commercial cellulases and demonstrated the capability of Y. lipolytica to convert cellulose into high-value organics such as α-ketoglutaric acid (KGA) in the SSF-IL process at relatively low temperature 28 °C and high pH 6.3. We showed that SSF-IL not only enhanced the enzymatic saccharification but also produced KGA up to 92 % of the maximum theoretical yield.
Similar content being viewed by others
References
Andreishcheva EN, Isakova EP, Sidorov NN, Abramova NB, Ushakova NA, Shaposhnikov GL, Soares MIM, Zvyagilskaya RA (1999) Adaptation to salt stress in a salt-tolerant strain of the yeast Yarrowia lipolytica. Biochemistry (Mosc) 64:1061–1067
Aurich A, Specht R, Müller RA, Stottmeister U, Yovkova V, Otto C, Holz M, Barth G, Heretsch P, Thomas FA (2012) Microbiologically produced carboxylic acids used as building blocks in organic synthesis. In: Wang X, Chen J, Quinn P (eds) Reprogramming microbial metabolic pathways. Springer, Dordrecht, pp 391–423
Beopoulos A, Cescut J, Haddouche R, Uribelarrea JL, Molina-Jouve C, Nicaud JM (2009) Yarrowia lipolytica as a model for bio-oil production. Prog Lipid Res 48:375–387
Boonvitthya N, Bozonnet S, Burapatana V, O’Donohue MJ, Chulalaksananukul W (2013) Comparison of the heterologous expression of Trichoderma reesei endoglucanase II and cellobiohydrolase II in the yeasts Pichia pastoris and Yarrowia lipolytica. Mol Biotechnol 54:158–169
Buschke N, Schäfer R, Becker J, Wittmann C (2013) Metabolic engineering of industrial platform microorganisms for biorefinery applications—optimization of substrate spectrum and process robustness by rational and evolutive strategies. Bioresour Technol 135:544–554
Chin RM, Fu X, Pai MY, Vergnes L, Hwang H, Deng G, Diep S, Lomenick B, Meli VS, Monsalve GC, Hu E, Whelan SA, Wang JX, Jung G, Solis GM, Fazlollahi F, Kaweeteerawat C, Quach A, Nili M, Krall AS, Godwin HA, Chang HR, Faull KF, Guo F, Jiang M, Trauger SA, Saghatelian A, Braas D, Christofk HR, Clarke CF, Teitell MA, Petrascheck M, Reue K, Jung ME, Frand AR, Huang J (2014) The metabolite α-ketoglutarate extends lifespan by inhibiting ATP synthase and TOR. Nature 510:397–401
Chundawat SPS, Beckham GT, Himmel ME, Dale BE (2011) Deconstruction of lignocellulosic biomass to fuels and chemicals. Ann Rev Chem Biomol Eng 2:121–145
Coelho MAZ, Amaral PFF, Belo I (2010) Yarrowia lipolytica: an industrial workhorse. In: Mendez-Vilas A (ed) Current research, technology and education topics in applied microbiology and microbial biotechnology. Formatex Research Center, Badajoz, p 930e44
Da Costa Lopes A, João K, Morais A, Bogel-Łukasik E, Bogel-Łukasik R (2013) Ionic liquids as a tool for lignocellulosic biomass fractionation. Sustain Chem Process 1:1–31
Dobler D, Schmidts T, Klingenhofer I, Runkel F (2013) Ionic liquids as ingredients in topical drug delivery systems. Int J Pharm 441:620–627
Docherty KM, Kulpa CF (2005) Toxicity and antimicrobial activity of imidazolium and pyridinium ionic liquids. Green Chem 7:185–189
Engel P, Mladenov R, Wulfhorst H, Jäger G, Spieß AC (2010) Point by point analysis: how ionic liquid affects the enzymatic hydrolysis of native and modified cellulose. Green Chem 12:1959–1966
Epova E, Kovalyov L, Isakova E, Deryabina Y, Belyakova A, Zylkova M, Shevelev A (2012) Identification of proteins involved in pH adaptation in extremophile yeast Yarrowia lipolytica. In: Heazlewood JL, Petzold CJ (eds) Proteomic applications in biology. Intech Europe, Rijeka, pp 209–224
Eyal AM, Lehnhardt WF (1998) Production and recovery of organic acids. US Patent and Trademark Office, Washington, DC, US Patent No. 5,766,439
Finogenova TV, Morgunov IG, Kamzolova SV, Chernyavskaya OG (2005) Organic acid production by the yeast Yarrowia lipolytica: a review of prospects. Appl Biochem Microbiol 41:418–425
Garcia MT, Gathergood N, Scammells PJ (2005) Biodegradable ionic liquids part II: effect of the anion and toxicology. Green Chem 7:9–14
Ghose TK (1987) Measurement of cellulase activities. Pure Appl Chem 59:257–268
Groenewald M, Boekhout T, Neuveglise C, Gaillardin C, van Dijck PWM, Wyss M (2014) Yarrowia lipolytica: safety assessment of an oleaginous yeast with a great industrial potential. Crit Rev Microbiol 40:187–206
Haghighi Mood S, Hossein Golfeshan A, Tabatabaei M, Salehi Jouzani G, Najafi GH, Gholami M, Ardjmand M (2013) Lignocellulosic biomass to bioethanol, a comprehensive review with a focus on pretreatment. Renew Sust Energ Rev 27:77–93
Himmel ME, Ding SY, Johnson DK, Adney WS, Nimlos MR, Brady JW, Foust TD (2007) Biomass recalcitrance: engineering plants and enzymes for biofuels production. Science 315:804–807
Kamzolova SV, Morgunov IG (2013) α-Ketoglutaric acid production from rapeseed oil by Yarrowia lipolytica yeast. Appl Microbiol Biotechnol 97:5517–5525
Kamzolova SV, Chiglintseva MN, Lunina JN, Morgunov IG (2012) α-Ketoglutaric acid production by Yarrowia lipolytica and its regulation. Appl Microbiol Biotechnol 96:783–791
Karasu-Yalcin S, Bozdemir MT, Ozbas ZY (2010) Effects of different fermentation conditions on growth and citric acid production kinetics of two Yarrowia lipolytica strains. Chem Biochem Eng Q 24:347–360
Khudyakov JI, D’haeseleer P, Borglin SE, DeAngelis KM, Woo H, Lindquist EA, Hazen TC, Simmons BA, Thelen MP (2012) Global transcriptome response to ionic liquid by a tropical rain forest soil bacterium, Enterobacter lignolyticus. Proc Natl Acad Sci U S A 109:E2173–E2182
Klein-Marcuschamer D, Simmons BA, Blanch HW (2011) Techno-economic analysis of a lignocellulosic ethanol biorefinery with ionic liquid pre-treatment. Biofuels Bioprod Bioref 5:562–569
Konda NM, Shi J, Singh S, Blanch HW, Simmons BA, Klein-Marcuschamer D (2014) Understanding cost drivers and economic potential of two variants of ionic liquid pretreatment for cellulosic biofuel production. Biotechnol Biofuels 7:86
Lan TQ, Lou H, Zhu JY (2013) Enzymatic saccharification of lignocelluloses should be conducted at elevated pH 5.2–6.2. BioEnerg Res 6:476–485
Lee SH, Doherty TV, Linhardt RJ, Dordick JS (2009) Ionic liquid‐mediated selective extraction of lignin from wood leading to enhanced enzymatic cellulose hydrolysis. Biotechnol Bioeng 102:1368–1376
Li C, Knierim B, Manisseri C, Arora R, Scheller HV, Auer M, Vogel KP, Simmons BA, Singh S (2010) Comparison of dilute acid and ionic liquid pretreatment of switchgrass: biomass recalcitrance, delignification and enzymatic saccharification. Bioresour Technol 101:4900–4906
Li C, Cheng G, Balan V, Kent MS, Ong M, Chundawat SPS, Ld S, Melnichenko YB, Dale BE, Simmons BA, Singh S (2011) Influence of physico-chemical changes on enzymatic digestibility of ionic liquid and AFEX pretreated corn stover. Bioresour Technol 102:6928–6936
Li C, Sun L, Simmons B, Singh S (2013) Comparing the recalcitrance of eucalyptus, pine, and switchgrass using ionic liquid and dilute acid pretreatments. BioEnerg Res 6:14–23
Liu C-Z, Wang F, Stiles AR, Guo C (2012) Ionic liquids for biofuel production: opportunities and challenges. Appl Energy 92:406–414
Liu XH, Li L, Yu ST, Zhu NN, Su ZL, Liu SW, Liu FS, Xie CX, Zhang BQ, Zhang CG (2014) Study on enzymatic degradation of cornstalk in ionic liquid. Catal Lett 144:229–234
Lynd LR, Weimer PJ, van Zyl WH, Pretorius IS (2002) Microbial cellulose utilization: fundamentals and biotechnology. Microbiol Mol Biol Rev 66:506–577
Mora‐Pale M, Meli L, Doherty TV, Linhardt RJ, Dordick JS (2011) Room temperature ionic liquids as emerging solvents for the pretreatment of lignocellulosic biomass. Biotechnol Bioeng 108:1229–1245
Morgunov IG, Kamzolova SV, Samoilenko VA (2013) Enhanced α-ketoglutaric acid production and recovery in Yarrowia lipolytica yeast by effective pH controlling. Appl Microbiol Biotechnol 97:8711–8718
Mosier N, Wyman C, Dale B, Elander R, Lee YY, Holtzapple M, Ladisch M (2005) Features of promising technologies for pretreatment of lignocellulosic biomass. Bioresour Technol 96:673–686
Nakashima K, Yamaguchi K, Taniguchi N, Arai S, Yamada R, Katahira S, Ishida N, Takahashi H, Ogino C, Kondo A (2011) Direct bioethanol production from cellulose by the combination of cellulase-displaying yeast and ionic liquid pretreatment. Green Chem 13:2948–2953
Otto C, Yovkova V, Barth G (2011) Overproduction and secretion of α-ketoglutaric acid by microorganisms. Appl Microbiol Biotechnol 92:689–695
Ouellet M, Datta S, Dibble DC, Tamrakar PR, Benke PI, Li CL, Singh S, Sale KL, Adams PD, Keasling JD, Simmons BA, Holmes BM, Mukhopadhyay A (2011) Impact of ionic liquid pretreated plant biomass on Saccharomyces cerevisiae growth and biofuel production. Green Chem 13:2743–2749
Park S, Kazlauskas RJ (2003) Biocatalysis in ionic liquids—advantages beyond green technology. Curr Opin Biotechnol 14:432–437
Park JI, Steen EJ, Burd H, Evans SS, Redding-Johnson AM, Batth T, Benke PI, D’Haeseleer P, Sun N, Sale KL, Keasling JD, Lee TS, Petzold CJ, Mukhopadhyay A, Singer SW, Simmons BA, Gladden JM (2012) A thermophilic ionic liquid-tolerant cellulase cocktail for the production of cellulosic biofuels. PLoS One 7:e37010
Peralta-Yahya PP, Zhang F, del Cardayre SB, Keasling JD (2012) Microbial engineering for the production of advanced biofuels. Nature 488:320–328
Ruegg TL, Kim EM, Simmons BA, Keasling JD, Singer SW, Soon Lee T, Thelen MP (2014) An auto-inducible mechanism for ionic liquid resistance in microbial biofuel production. Nat Commun 5:3490
Santos AG, Ribeiro BD, Alviano DS, Coelho M (2014) Toxicity of ionic liquids toward microorganisms interesting to the food industry. RSC Adv 4:37157–37163
Shi J, Gladden JM, Sathitsuksanoh N, Kambam P, Sandoval L, Mitra D, Zhang S, George A, Singer SW, Simmons BA, Singh S (2013) One-pot ionic liquid pretreatment and saccharification of switchgrass. Green Chem 15:2579–2589
Shill K, Padmanabhan S, Xin Q, Prausnitz JM, Clark DS, Blanch HW (2011) Ionic liquid pretreatment of cellulosic biomass: enzymatic hydrolysis and ionic liquid recycle. Biotechnol Bioeng 108:511–520
Shu G, Yang H, Chen H, Yang Z (2013) Research on extraction and characterization of cellulase from commercial enzyme preparation. Adv J Food Sci Technol 5:839–842
Stephanopoulos G (2007) Challenges in engineering microbes for biofuels production. Science 315:801–804
Stottmeister U, Aurich A, Wilde H, Andersch J, Schmidt S, Sicker D (2005) White biotechnology for green chemistry: fermentative 2-oxocarboxylic acids as novel building blocks for subsequent chemical syntheses. J Ind Microbiol Biotechnol 32:651–664
Thanh HT, Beney L, Simonin H, Nguyen TXS, Gervais P, Belin JM, Husson F (2007) Toxicity of fatty acid hydroperoxides towards Yarrowia lipolytica: implication of their membrane fluidizing action. Biochim Biophys Acta 1768:2256–2262
Thi PTP, Cho CW, Yun YS (2010) Environmental fate and toxicity of ionic liquids: a review. Water Res 44:352–372
Trinh CT, Unrean P, Srienc F (2008) Minimal Escherichia coli cell for the most efficient production of ethanol from hexoses and pentose. Appl Environ Microbiol 74:3634–3643
Turk M, Abramović Z, Plemenitaš A, Gunde‐Cimerman N (2007) Salt stress and plasma‐membrane fluidity in selected extremophilic yeasts and yeast‐like fungi. FEMS Yeast Res 7:550–557
Turner MB, Spear SK, Huddleston JG, Holbrey JD, Rogers RD (2003) Ionic liquid salt-induced inactivation and unfolding of cellulase from Trichoderma reesei. Green Chem 5:443–447
Van Rantwijk F, Sheldon RA (2007) Biocatalysis in ionic liquids. Chem Rev 107:2757–2785
Varanasi P, Singh P, Auer M, Adams PD, Simmons BA, Singh S (2013) Survey of renewable chemicals produced from lignocellulosic biomass during ionic liquid pretreatment. Biotechnol Biofuels 6:14
Wang JH, Hung WP, Tsai SH (2011a) High efficiency transformation by electroporation of Yarrowia lipolytica. J Microbiol 49:469–472
Wang Y, Radosevich M, Hayes D, Labbe N (2011b) Compatible ionic liquid-cellulases system for hydrolysis of lignocellulosic biomass. Biotechnol Bioeng 108:1042–1048
Yamane T, Sakai H, Nagahama K, Ogawa T, Matsuoka M (2008) Dissection of centromeric DNA from yeast Yarrowia lipolytica and identification of protein-binding site required for plasmid transmission. J Biosci Bioeng 105:571–578
Yang B, Wyman CE (2008) Pretreatment: the key to unlocking low‐cost cellulosic ethanol. Biofuels Bioprod Bioref 2:26–40
Zhou JW, Zhou HY, Du GC, Liu LM, Chen JA (2010) Screening of a thiamine-auxotrophic yeast for α-ketoglutaric acid overproduction. Lett Appl Microbiol 51:264–271
Acknowledgments
This research was financially supported by the Institute of a Secure and Sustainable Environment/ Sustainable Energy and Education Research Center (CTT and NL), the laboratory start-up fund (CTT) at the University of Tennessee, Knoxville, and the National Science Foundation, NSF-CBET# 1360867 (CTT).
Conflict of interest
The authors claim no conflict of interest.
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
ESM 1
(PDF 102 kb)
Rights and permissions
About this article
Cite this article
Ryu, S., Labbé, N. & Trinh, C.T. Simultaneous saccharification and fermentation of cellulose in ionic liquid for efficient production of α-ketoglutaric acid by Yarrowia lipolytica . Appl Microbiol Biotechnol 99, 4237–4244 (2015). https://doi.org/10.1007/s00253-015-6521-5
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-015-6521-5