Abstract
The fermentative and respiratory metabolism of Pichia stipitis wild-type strain CBS 5774 and the derived auxotrophic transformation recipient PJH53 trp5-10 his3-1 were examined in differentially oxygenated glucose cultures in the hermetically sealed Sensomat system. There was a good agreement of the kinetics of gas metabolism, growth, ethanol formation and glucose utilisation, proving the suitability of the Sensomat system for rapid and inexpensive investigation of strains and mutants for their respiratory and fermentative metabolism. Our study revealed respiro-fermentative growth by the wild-type strain, although the cultures were not oxygen-limited. The induction of respiro-fermentative behaviour was obviously due to the decrease in oxygen tension but not falling below a threshold of oxygen tension. The responses differed depending on the velocity of the decrease in oxygen tension. At high oxygenation (slow decrease in oxygen tension), ethanol production was induced but glucose uptake was not influenced. At low oxygenation, glucose uptake and ethanol formation increased during the first hours of cultivation. The transformation recipient PJH53 most probably carries a mutation that influences the response to a slow decrease in oxygen tension, since almost no ethanol formation was found under these conditions.
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Ausubel FM, Brent R, Kington RE, Moore DD, Seidman JG, Smith JA, Struhl K (1991) Current protocols in molecular biology. Wiley, New York
Bergmeyer HU (1974) Methoden der enzymatischen Analyse. Chemie, Weinheim
Bergter F (1983) Wachstum von Mikroorganismen. Experimente und Modelle. Chemie, Weinheim
Bracken CP, Whitelaw ML, Peet DJ (2003) The hypoxia-inducible factors: key transcriptional regulators of hypoxic responses. Cell Mol Life Sci 60:1376–1393
Cho JY, Jeffries TW (1998) Pichia stipitis genes for alcohol dehydrogenase with fermentative and respiratory functions. Appl Environ Microbiol 64:1350–1358
Cho JY, Jeffries TW (1999) Transcriptional control of ADH genes in the xylose-fermenting yeast Pichia stipitis. Appl Environ Microbiol 65:2363–2368
Dellweg H, Rizzi M, Methner H, Debus D (1984) Xylose fermentation by yeasts 3. Comparison of Pachysolen tannophilus and Pichia stipitis. Biotechnol Lett 6:395–400
Dijken JP van, Scheffers WA (1986) Redox balances in the metabolism of sugars by yeasts. FEMS Microbiol Rev 32:199–224
Elsworth R, Williams V, Harris-Smith R (1957) The effect of oxygen supply on the rate of growth in Aerobacter cloacae. J Appl Chem 7:269–274
Fiaux J, Cakar ZP, Sonderegger M, Wüthrich K, Szyperski T, Sauer U (2003) Metabolic-flux profiling of the yeasts Saccharomyces cerevisiae and Pichia stipitis. Eukaryot Cell 2:170–180
Fluthgraf S, Kirchhoff A, Debye J, Passoth V, Klinner U (2003) Non-conventional yeasts in genetics, biochemistry and biotechnology. Springer, Berlin Heidelberg New York, pp 229–223
Gancedo C, Serrano R (1989) Energy-yielding metabolism. In: Rose AH, Harrison JS (eds) The yeasts, vol 3. Academic, London, pp 205–259
Hagedorn J (1990) Isolierung und Charakterisierung von Mutanten im Xylosestoffwechsel und Entwicklung eines Transformationssystems für die Hefe Pichia stipitis. PhD thesis, University of Düsseldorf, Düsseldorf
Hensing MCM, Rouwenhorst RJ, Heijnen JJ, Dijken JP van, Pronk JT (1995) Physiological and technological aspects of large-scale heterologous-protein production with yeasts. Antonie van Leeuwenhoek 67:261–279
Jones EW, Fink GR (1982) Regulation of amino acid and nucleotide biosysnthesis in yeast. In: Strathern JN, Jones EW, Broach JR (eds) The molecular biology of the yeast Saccharomyces. Metabolism and gene expression. Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y., pp 181–299
Lighthelm ME, Prior BA, du Preez JC (1988) The oxygen requirements of yeasts for the fermentation of d-xylose and d-glucose to ethanol. Appl Microbiol Biotechnol 28:63–68
Mergler M, Klinner U (2001) Cell density-correlated induction of pyruvate decarboxylase under aerobic conditions in the yeast Pichia stipitis. Acta Biol Hung 52:265–269
Melake T, Passoth V, Klinner U (1996) Characterization of the genetic system of the xylose-fermenting yeast Pichia stipitis. Curr Microbiol 33:237–242
Morosoli R, Zalce E, Durand S (1993) Secretion of a Cryptococcus albidus xylanase in Pichia stipitis resulting in a xylan fermenting transformant. Curr Genet 24:94–99
Passoth V, Hahn-Hägerdal B (2000) Production of a heterologous endo-1,4-beta-xylanase in the yeast Pichia stipitis with an O2-regulated promoter. Enzyme Microb Technol 26:781–784
Passoth V, Zimmermann M, Klinner U (1996) Peculiarities of the regulation of fermentation and respiration in the Crabtree-negative, xylose-fermenting yeast Pichia stipitis. Appl Biochem Biotechnol 57/58:201–212
Passoth V, Schäfer B, Liebel B, Weierstall T, Klinner U (1998) Molecular cloning of alcohol dehydrogenase genes of the yeast Pichia stipitis and identification of the fermentative ADH. Yeast 14:1311–1325
Passoth V, Cohn M, Schäfer B, Hahn-Hägerdal B, Klinner U (2003) Analysis of the hypoxia-induced ADH2 promoter of the respiratory yeast Pichia stipitis reveals a new mechanism of hypoxia sensing of oxygen limitation in yeast. Yeast 20:39–51
Petrik M, Käppeli O, Fiechter A (1983) An expanded concept for the glucose effect in the yeast Saccharomyces-uvarum— involvement of short-term and long-term regulation. J Gen Microbiol 129:43–49
Piontek M, Hagedorn J, Hollenberg CP, Gellissen G, Strasser AWM (1998) Two novel gene expression systems based on the yeasts Schwanniomyces occidentalis and Pichia stipitis. Appl Microbiol Biotechnol 50:331–338
du Preez JC (1994) Process parameters and environmental factors affecting d-xylose fermentation by yeasts. Enzyme Microb Technol 16:944–956
Pronk JT (2002) Auxotrophic yeast strains in fundamental and applied research. Appl Environ Microbiol 68:2095–2100
Skoog K, Jeppsson H, Hahn-Hägerdal B (1992) The effect of oxygenation on glucose fermentation with Pichia stipitis. Appl Biochem Biotechnol 34/35:369–375
Stuehr DJ (1999) Mammalian nitric oxide synthases. Biochim Biophys Acta Bioenergetics 1411:217–230
Zitomer RS, Carrico P, Deckert J (1997) Regulation of hypoxic gene expression in yeast. Kidney Int 51:507–513
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This work was supported by the Deutsche Forschungsgemeinschaft. We thank Ulrike Schmitt for linguistic advice.
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Klinner, U., Fluthgraf, S., Freese, S. et al. Aerobic induction of respiro-fermentative growth by decreasing oxygen tensions in the respiratory yeast Pichia stipitis. Appl Microbiol Biotechnol 67, 247–253 (2005). https://doi.org/10.1007/s00253-004-1746-8
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DOI: https://doi.org/10.1007/s00253-004-1746-8