Abstract
Objectives
Development of an open-loop fed-batch protocol for highly reproducible fermentation of fission yeast that starts from batch cultures instead of glucose-limited aerobic chemostat cultures.
Results
A new strategy was employed that consists of an exponential feeding phase followed by a starvation period and then a linear feeding phase. A comparison of several independent fed-batch fermentations of a recombinant fission yeast strain showed that while during the initial phase process parameters such as glucose consumption and CO2 evolution varied considerably as expected, they were much more uniform during the third phase. For instance, the normalized standard deviation of glucose consumption was thirty times higher during the exponential feeding phase of the fermentation that during the linear feeding phase.
Conclusion
These data demonstrate the usefulness of the proposed strategy. It is expected that by variation of only two parameters (the total amount of glucose fed in the initial phase and the time frame of the starvation phase) the protocol can easily be adapted to other microbes.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Benito S (2019) The impacts of Schizosaccharomyces on winemaking. Appl Microbiol Biotechnol 103:4291–4312. https://doi.org/10.1007/s00253-019-09827-7
Dragan CA, Buchheit D, Bischoff D, Ebner T, Bureik M (2010) Glucuronide production by whole-cell biotransformation using genetically engineered fission yeast Schizosaccharomyces pombe. Drug Metab Dispos 38:509–515
Durairaj P et al (2019) Functional expression and activity screening of all human cytochrome P450 enzymes in fission yeast. FEBS Lett 593:1372–1380. https://doi.org/10.1002/1873-3468.13441
Forsburg SL, Rhind N (2006) Basic methods for fission yeast. Yeast 23:173–183. https://doi.org/10.1002/yea.1347
Giga-Hama Y, Kumagai H (1997) Foreign gene expression in fission yeast Schizosaccharomyces pombe. Springer, Berlin
Goldrick S, Lee K, Spencer C, Holmes W, Kuiper M, Turner R, Farid SS (2018) On-line control of glucose concentration in high-yielding mammalian cell cultures enabled through oxygen transfer rate measurements. Biotechnol J. https://doi.org/10.1002/biot.201700607
Ikeda S, Nikaido K, Araki K, Yoshitake A, Kumagai H, Isoai A (2004) Production of recombinant human lysosomal acid lipase in Schizosaccharomyces pombe: development of a fed-batch fermentation and purification process. J Biosci Bioeng 98:366–373
Jansen ML, Krook DJ, De Graaf K, van Dijken JP, Pronk JT, de Winde JH (2006) Physiological characterization and fed-batch production of an extracellular maltase of Schizosaccharomyces pombe CBS 356. FEMS Yeast Res 6:888–901
Jenzsch M, Gnoth S, Beck M, Kleinschmidt M, Simutis R, Lubbert A (2006) Open-loop control of the biomass concentration within the growth phase of recombinant protein production processes. J Biotechnol 127:84–94. https://doi.org/10.1016/j.jbiotec.2006.06.004
Lindner P (1893) Schizosaccharomyces pombe n. sp., ein neuer Gährungserreger. Wochenschr Brau 10:1298–1300
Liu J et al (2018) Combined chemical and biotechnological production of 20betaOH-NorDHCMT, a long-term metabolite of oral-turinabol (DHCMT). J Inorg Biochem 183:165–171. https://doi.org/10.1016/j.jinorgbio.2018.02.020
Luttmann R et al (2012) Soft sensors in bioprocessing: a status report and recommendations. Biotechnol J 7:1040–1048. https://doi.org/10.1002/biot.201100506
Maundrell K (1990) nmt1 of fission yeast. A highly transcribed gene completely repressed by thiamine. J Biol Chem 265:10857–10864
Miller WL, Auchus RJ (2011) The molecular biology, biochemistry, and physiology of human steroidogenesis and its disorders. Endocr Rev 32:81–151
Naumann JM, Messinger J, Bureik M (2010) Human 20alpha-hydroxysteroid dehydrogenase (AKR1C1)-dependent biotransformation with recombinant fission yeast Schizosaccharomyces pombe. J Biotechnol 150:161–170
Naumann JM, Küttner G, Bureik M (2011a) Expression and secretion of a CB4-1 scFv-GFP fusion protein by fission yeast. Appl Biochem Biotechnol 163:80–89
Naumann JM, Zöllner A, Dragan CA, Messinger J, Adam J, Bureik M (2011b) Biotechnological production of 20-alpha-dihydrodydrogesterone at pilot scale. Appl Biochem Biotechnol 165:190–203
Neunzig I, Widjaja M, Peters FT, Maurer HH, Hehn A, Bourgaud F, Bureik M (2013) Coexpression of CPR from various origins enhances biotransformation activity of human CYPs in S. pombe. Appl Biochem Biotechnol 170:1751–1766
Peters FT, Dragan CA, Schwaninger AE, Sauer C, Zapp J, Bureik M, Maurer HH (2009) Use of fission yeast heterologously expressing human cytochrome P450 2B6 in biotechnological synthesis of the designer drug metabolite N-(1-phenylcyclohexyl)-2-hydroxyethanamine. Forensic Sci Int 184:69–73
Rolland D, Raymond F, Gauthier M, Fournier C, Charrier JP, Jolivet M, Dantigny P (2008) Strategies for improving production and purification of a recombinant protein: rP30 of Toxoplasma gondii expressed in the yeast Schizosaccharomyces pombe. J Chromatogr B 861:186–195. https://doi.org/10.1016/j.jchromb.2007.07.034
Schaepe S, Kuprijanov A, Simutis R, Lübbert A, Pohlscheidt M, Jenzsch M (2013) Batch-to-batch reproducibility of fermentation processes by robust operational design and control. Pharm Bioprocess 1:297–307
Srinivasan S, Feng SJ, Lin YH (2012) Dissolved carbon dioxide concentration profiles during very-high-gravity ethanol fermentation. Biochem Eng J 69:41–47. https://doi.org/10.1016/j.bej.2012.07.019
Stoll A et al (2019) Fine-mapping of the substrate specificity of human steroid 21-hydroxylase (CYP21A2). J Steroid Biochem Mol Biol 194:105446. https://doi.org/10.1016/j.jsbmb.2019.105446
Zhang D, Shrestha B, Niu W, Tian P, Tan T (2007) Phenotypes and fed-batch fermentation of ubiquinone-overproducing fission yeast using ppt1 gene. J Biotechnol 128:120–131
Zöllner A, Buchheit D, Meyer MR, Maurer HH, Peters FT, Bureik M (2010) Production of human phase 1 and 2 metabolites by whole-cell biotransformation with recombinant microbes. Bioanalysis 2:1277–1290
Zöllner A et al (2010) CYP21-catalyzed production of the long-term urinary metandienone metabolite 17beta-hydroxymethyl-17alpha-methyl-18-norandrosta-1,4,13-trien-3-one: a contribution to the fight against doping. Biol Chem 391:119–127
Supporting informations
Supplementary Fig. 1—Comparison of continuous carbon dioxide evolution (CO2%; bottom) and dissolved oxygen (DO %; top) measurements during three individual fed-batch cultivations of strain JMN8. Each color (purple, orange, magenta) represents the measurements of an individual fermentation run.
Supplementary Fig. 2—Fed-batch cultivation of strain CAD75 at pH 5.5. Total glucose fed (brown) was calculated continuously; samples were taken periodically and analyzed for glucose concentration (red) and biomass (blue) as indicated. Glucose consumption rates during different time intervals are shown in light purple.
Supplementary Fig. 3—Comparison of continuous carbon dioxide evolution (CO2 (%); bottom) and dissolved oxygen (DO; top) measurements during fed-batch cultivation of strain CAD75 at pH 5.5 (top row) and CO2 evolution rates (d([CO2])/dt; bottom row). The CO2 evolution rates are the first-order derivatives of the CO2 evolution profiles shown in the top row. The markings correspond to the respective regions in the CO2 evolution profiles.
Supplementary Fig. 4—Fed-batch cultivation of strain CAD75 at pH 3. Total glucose fed (brown) was calculated continuously; samples were taken periodically and analyzed for glucose concentration (red) and biomass (blue) as indicated. Glucose consumption rates during different time intervals are shown in light purple.
Supplementary Fig. 5—Comparison of continuous carbon dioxide evolution (CO2 (%); bottom) and dissolved oxygen (DO; top) measurements during fed-batch cultivation of strain CAD75 at pH 3 (top row) and CO2 evolution rates (d([CO2])/dt; bottom row). The CO2 evolution rates are the first-order derivatives of the CO2 evolution profiles shown in the top row. The markings correspond to the respective regions in the CO2 evolution profiles.
Author information
Authors and Affiliations
Corresponding author
Additional information
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Scomparin, A., Bureik, M. A convenient new method for reproducible fed-batch fermentation of fission yeast Schizosaccharomyces pombe. Biotechnol Lett 42, 937–943 (2020). https://doi.org/10.1007/s10529-020-02840-1
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10529-020-02840-1