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Received September 28, 2021
Accepted November 17, 2021
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Fluorescence-activated cell sorting-mediated directed evolution of Wickerhamomyces ciferrii for enhanced production of tetraacetyl phytosphingosine
Su-Bin Park1 2
Quynh-Giao Tran1 3
Ae Jin Ryu1 3
Jin-Ho Yun1 3
Kil Koang Kwon4
Yong Jae Lee1 2†
Hee-Sik Kim1 2†
1Cell Factory Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Korea 2Department of Environmental Biotechnology, KRIBB School of Biotechnology, University of Science and Technology (UST), Daejeon 34113, Korea 3, Korea 4Synthethic Biology & Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Korea
Korean Journal of Chemical Engineering, April 2022, 39(4), 1004-1010(7), 10.1007/s11814-021-1017-8
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Abstract
Ceramides are a major lipid class known to play an essential role in maintaining skin function. Thus, efforts have been made to produce ceramides and ceramide precursors in large quantities for industrial applications. The yeast Wickerhamomyces ciferrii, a natural producer of the ceramide precursor tetraacetyl phytosphingosine (TAPS), has been isolated and engineered through various mutagenesis approaches aiming to enhance TAPS production. Herein, a highthroughput screening platform for isolating W. ciferrii mutants with improved TAPS production is described. A fluorescence- mediated reporter system that allows initial quantification of TAPS content in yeast cells based on BODIPY staining was developed. The optimal concentration of BODIPY for monitoring intracellular TAPS levels in W. ciferrii was 400 μg/L, as shown by a linear correlation between the actual TAPS levels and mean fluorescence intensities. Fluorescence- activated cell sorting was used for isolating high TAPS-producing strains from an ethyl methanesulfonateinduced mutant library. After several rounds of sorting, mutants exhibiting a high-TAPS phenotype were isolated, and the M40 strain with the highest TAPS titer was chosen for large-scale cultivation. The influence of different carbon sources for optimizing TAPS production was also evaluated using the M40 strain. A maximum production yield of 5.114 g/L of ceramide precursors, including TAPS and triacetyl phytosphingosine, was achieved with the supplementation of molasses. This novel platform enables rapid screening of high TAPS-producing strains using the common dye BODIPY and can be easily extended for the development of mutants with high productivity of ceramide precursors in yeast and other microorganisms.
Keywords
References
Bouwstra JA, Ponec M, Biochim. Biophys. Acta. Biomembr., 1758, 2080
Elias PM, Williams ML, Feingold KR, Clin. Dermatol., 30, 311 (2012)
Li Q, Fang H, Dang E, Wang G, J. Dermatol. Sci., 97, 2 (2020)
Sano S, Dermatol. Sin., 33, 64 (2015)
Sajna KV, Gottumukkala LD, Sukumaran RK, Pandey A, in Industrial biorefineries & white biotechnology, Pandey A, Höfer R, Taherzadeh M, Nampoothiri KM, Larroche C Eds., Elsevier, Amsterdam (2015).
Gault CR, Obeid LM, Hannun YA, Adv. Exp. Med. Biol., 688, 1 (2010)
Liotta D, Merrill AH, Canada Patent, CA1340549C (1999).
Nugent TC, Hudlicky T, J. Org. Chem., 63, 510 (1998)
Rochlin E, CA Patent, 2,373,286C (2010).
Zhang L, Hellgren LI, Xu X, J. Biotechnol., 123, 93 (2006)
Stodola FH, Wickerham LJ, J. Biol. Chem., 235, 2584 (1960)
Wickerham LJ, Stodola FH, J. Bacteriol., 80, 484 (1960)
Maister HG, Rogovin SP, Stodola FH, Wickerham LJ, Appl. Microbiol., 10, 401 (1962)
Kreger-van Rij NJW, The yeasts: A taxonomic study (3rd ed. rev.), Elsevier, Amsterdam (2013).
Schorsch C, Köhler T, Andrea H, Boles E, Metab. Eng., 14, 172 (2012)
Arora N, Yen HW, Philippidis GP, Sustainability, 12, 5125 (2020)
Kacmar J, Carlson R, Balogh SJ, Srienc F, Cytometry A, 69A, 27 (2006)
Cooper MS, Hardin WR, Petersen TW, Cattolico RA, J. Biosci. Bioeng., 109, 198 (2010)
Spencer JFT, Spencer DM, in Yeast protocols: Methods in cell and molecular biology, Evans IH Eds., Springer Nature, Switzerland (1996).
Martinet W, Maras M, Saelens X, Jou WM, Contreras R, Biotechnol. Lett., 20, 1171 (1998)
Panagiotou V, Love KR, Jiang B, Nett J, Stadheim T, Love JC, Appl. Environ. Microbiol., 77, 3154 (2011)
Ryu AJ, Kang NK, Jeon S, Hur DH, Lee EM, Lee DY, Jeong BR, Chang YK, Jeong KJ, Biotechnol. Biofuels, 13, 1 (2020)
Kang NK, Jeon S, Kwon S, Koh HG, Shin SE, Lee B, Choi GG, Yang JW, Jeong BR, Chang YK, Biotechnol. Biofuels, 8, 1 (2015)
Daugherty PS, Iverson BL, Georgiou G, J. Immunol. Methods, 243, 211 (2000)
Winston F, Curr. Protoc. Mol. Biol., 82, 13.3B.1 (2008)
Choi JY, Hwang HJ, Cho WY, Choi JI, Lee PC, Front. Bioeng. Biotechnol., 9, 662979 (2021)
Bekatorou A, Psarianos C, Koutinas AA, Food Technol. Biotechnol., 44, 407 (2006)
Clarke MA, in Encyclopedia of food sciences and nutrition (2nd ed.), Caballero B, Finglas PM, Trugo LC Eds., Academic Press, Oxford (2003).
Ometto AR, Hauschild MZ, Roma WNL, Int. J. Life Cycle Assess., 14, 236 (2009)
Ong KL, Kaur G, Pensupa N, Uisan K, Lin CSK, Bioresour. Technol., 248, 100 (2018)
Elias PM, Williams ML, Feingold KR, Clin. Dermatol., 30, 311 (2012)
Li Q, Fang H, Dang E, Wang G, J. Dermatol. Sci., 97, 2 (2020)
Sano S, Dermatol. Sin., 33, 64 (2015)
Sajna KV, Gottumukkala LD, Sukumaran RK, Pandey A, in Industrial biorefineries & white biotechnology, Pandey A, Höfer R, Taherzadeh M, Nampoothiri KM, Larroche C Eds., Elsevier, Amsterdam (2015).
Gault CR, Obeid LM, Hannun YA, Adv. Exp. Med. Biol., 688, 1 (2010)
Liotta D, Merrill AH, Canada Patent, CA1340549C (1999).
Nugent TC, Hudlicky T, J. Org. Chem., 63, 510 (1998)
Rochlin E, CA Patent, 2,373,286C (2010).
Zhang L, Hellgren LI, Xu X, J. Biotechnol., 123, 93 (2006)
Stodola FH, Wickerham LJ, J. Biol. Chem., 235, 2584 (1960)
Wickerham LJ, Stodola FH, J. Bacteriol., 80, 484 (1960)
Maister HG, Rogovin SP, Stodola FH, Wickerham LJ, Appl. Microbiol., 10, 401 (1962)
Kreger-van Rij NJW, The yeasts: A taxonomic study (3rd ed. rev.), Elsevier, Amsterdam (2013).
Schorsch C, Köhler T, Andrea H, Boles E, Metab. Eng., 14, 172 (2012)
Arora N, Yen HW, Philippidis GP, Sustainability, 12, 5125 (2020)
Kacmar J, Carlson R, Balogh SJ, Srienc F, Cytometry A, 69A, 27 (2006)
Cooper MS, Hardin WR, Petersen TW, Cattolico RA, J. Biosci. Bioeng., 109, 198 (2010)
Spencer JFT, Spencer DM, in Yeast protocols: Methods in cell and molecular biology, Evans IH Eds., Springer Nature, Switzerland (1996).
Martinet W, Maras M, Saelens X, Jou WM, Contreras R, Biotechnol. Lett., 20, 1171 (1998)
Panagiotou V, Love KR, Jiang B, Nett J, Stadheim T, Love JC, Appl. Environ. Microbiol., 77, 3154 (2011)
Ryu AJ, Kang NK, Jeon S, Hur DH, Lee EM, Lee DY, Jeong BR, Chang YK, Jeong KJ, Biotechnol. Biofuels, 13, 1 (2020)
Kang NK, Jeon S, Kwon S, Koh HG, Shin SE, Lee B, Choi GG, Yang JW, Jeong BR, Chang YK, Biotechnol. Biofuels, 8, 1 (2015)
Daugherty PS, Iverson BL, Georgiou G, J. Immunol. Methods, 243, 211 (2000)
Winston F, Curr. Protoc. Mol. Biol., 82, 13.3B.1 (2008)
Choi JY, Hwang HJ, Cho WY, Choi JI, Lee PC, Front. Bioeng. Biotechnol., 9, 662979 (2021)
Bekatorou A, Psarianos C, Koutinas AA, Food Technol. Biotechnol., 44, 407 (2006)
Clarke MA, in Encyclopedia of food sciences and nutrition (2nd ed.), Caballero B, Finglas PM, Trugo LC Eds., Academic Press, Oxford (2003).
Ometto AR, Hauschild MZ, Roma WNL, Int. J. Life Cycle Assess., 14, 236 (2009)
Ong KL, Kaur G, Pensupa N, Uisan K, Lin CSK, Bioresour. Technol., 248, 100 (2018)
