Abstract
This review describes progress using the yeast Saccharomyces cerevisiae as a model organism for the fast and efficient analysis of genes and enzyme activities involved in the lipid biosynthetic pathways of several donor organisms. Furthermore, we assess the impact of baker′s yeast on the production of novel, high-value lipid compounds. Yeast can be genetically modified to produce selected substances in relatively high amounts. A major advantage in choosing yeast as an object for metabolic engineering is the fact that the lipid pathways in this organism have been described in detail and are well characterized. We focus on the de novo production of three major families of lipid products. These are: (1) sterols, providing some previously known and some novel applications as examples of the lipid pathway enhancement that occurs naturally in yeast, (2) the reconstitution of the biosynthetic pathway of steroid hormones and (3) the biosynthesis of polyunsaturated fatty acids, leading to the biosynthesis of different omega-3 and omega-6 fatty acids which do not occur naturally in yeast. We utilize the current knowledge and point out perspectives and problems for future biotechnological applications in the field of lipid compounds.
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Aki T, Shimada Y, Inagaki K, Higashimoto H, Kawamoto S, Shigeta S, Ono K, Suzuki O (1999) Molecular cloning and functional characterization of rat delta-6 fatty acid desaturase. Biochem Biophys Res Commun 255:575–579
Alvarez HM, Steinbuchel A (2002) Triacylglycerols in prokaryotic microorganisms. Appl Microbiol Biotechnol 60:367–376
Arnezeder C, Hampel WA (1990) Influence of growth rate on the accumulation of ergosterol in yeast-cells. Biotechnol Lett 12:277–282
Bach AC, Ferezou J, Frey A (1996) Phospholipid-rich particles in commercial parenteral fat emulsions. An overview. Prog Lipid Res 35:133–153
Bagga D, Anders KH, Wang HJ, Glaspy JA (2002) Long-chain n-3-to-n-6 polyunsaturated fatty acid ratios in breast adipose tissue from women with and without breast cancer. Nutr Cancer 42:180–185
Bammert GF, Fostel JM (2000) Genome-wide expression patterns in Saccharomyces cerevisiae: comparison of drug treatments and genetic alterations affecting biosynthesis of ergosterol. Antimicrob Agents Chemother 44:1255–1265
Bard M, Lees ND, Turi T, Craft D, Cofrin L, Barbuch R, Koegel C, Loper JC (1993) Sterol synthesis and viability of erg11 (cytochrome P450 lanosterol demethylase) mutations in Saccharomyces cerevisiae and Candida albicans. Lipids 28:963–967
Baudry K, Swain E, Rahier A, Germann M, Batta A, Rondet S, Mandala S, Henry K, Tint GS, Edlind T, Kurtz M, Nickels JT Jr (2001) The effect of the erg26-1 mutation on the regulation of lipid metabolism in Saccharomyces cerevisiae. J Biol Chem 276:12702–12711
Beaudoin F, Michaelson LV, Hey SJ, Lewis MJ, Shewry PR, Sayanova O, Napier JA (2000) Heterologous reconstitution in yeast of the polyunsaturated fatty acid biosynthetic pathway. Proc Natl Acad Sci USA 97:6421–6426
Behalova B, Hozak P, Blahova M, Sillinger V (1992) Effect of nitrogen limitation and sporulation on sterol and lipid formation in Saccharomyces cerevisiae. Folia Microbiol (Praha) 37:442–449
Bloch KE (1983) Sterol structure and membrane function. Crit Rev Biochem 14:47–92
Broun P, Gettner S, Somerville C (1999) Genetic engineering of plant lipids. Annu Rev Nutr 19:197–216
Byskov AG, Andersen CY, Leonardsen L, Baltsen M (1999) Meiosis activating sterols (MAS) and fertility in mammals and man. J Exp Zool 285:237–242
Casey WM, Keesler GA, Parks LW (1992) Regulation of partitioned sterol biosynthesis in Saccharomyces cerevisiae. J Bacteriol 174:7283–7288
Cauet G, Degryse E, Ledoux C, Spagnoli R, Achstetter T (1999) Pregnenolone esterification in Saccharomyces cerevisiae. A potential detoxification mechanism. Eur J Biochem 261:317–324
Ciesarová Z, Sajbidor J, Smogrovicova, D, Bafrncova P (1996) Effect of ethanol on fermentation and lipid composition in Saccharomyces cerevisiae. Food Biotechnol 10:1–12
Covello PS, Reed DW (1996) Functional expression of the extraplastidial Arabidopsis thaliana oleate desaturase gene (FAD2) in Saccharomyces cerevisiae. Plant Physiol 111:223–226
Daum G, Lees ND, Bard M, Dickson R (1998) Biochemistry, cell biology and molecular biology of lipids of Saccharomyces cerevisiae. Yeast 14:1471–1510
Degryse E, Cauet G, Spagnoli R, Achstetter T (1999) Pregnenolone metabolized to 17alpha-hydroxyprogesterone in yeast: biochemical analysis of a metabolic pathway. J Steroid Biochem Mol Biol 71:239–246
Donald KA, Hampton RY, Fritz IB (1997) Effects of overproduction of the catalytic domain of 3-hydroxy-3-methylglutaryl coenzyme A reductase on squalene synthesis in Saccharomyces cerevisiae. Appl Environ Microbiol 63:3341–3344
Duport C, Spagnoli R, Degryse E, Pompon D (1998) Self-sufficient biosynthesis of pregnenolone and progesterone in engineered yeast. Nat Biotechnol 16:186–189
Duport C, Schoepp B, Chatelain E, Spagnoli R, Dumas B, Pompon D (2003) Critical role of the plasma membrane for expression of mammalian mitochondrial side chain cleavage activity in yeast. Eur J Biochem 270:1502–1514
Dyer JM, Chapital DC, Kuan JW, Mullen RT, Pepperman AB (2002) Metabolic engineering of Saccharomyces cerevisiae for production of novel lipid compounds. Appl Microbiol Biotechnol 59:224–230
Fannon SA, Vidaver RM, Marts SA (2001) An abridged history of sex steroid hormone receptor action. J Appl Physiol 91:1854–1859
Ford G, Ellis EM (2002) Characterization of Ypr1p from Saccharomyces cerevisiae as a 2-methylbutyraldehyde reductase. Yeast 19:1087–1096
Gao H, Tan TW (2001) Analysis of parameters for optimum of ergosterol fermentation. Sheng Wu Gong Cheng Xue Bao 17:693–697
Gardner RG, Hampton RY (1999) A highly conserved signal controls degradation of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase in eukaryotes. J Biol Chem 274:31671–31678
Gibson RA, Makrides M (1998) The role of long chain polyunsaturated fatty acids (LCPUFA) in neonatal nutrition. Acta Paediatr 87:1017–1022
Gill I, Valivety R (1997) Polyunsaturated fatty acids, part 1: occurrence, biological activities and applications. Trends Biotechnol 15:401–409
Grabinska K, Palamarczyk G (2002) Dolichol biosynthesis in the yeast Saccharomyces cerevisiae: an insight into the regulatory role of farnesyl diphosphate synthase. FEMS Yeast Res 2:259–265
Grunler J, Ericsson J, Dallner G (1994) Branch-point reactions in the biosynthesis of cholesterol, dolichol, ubiquinone and prenylated proteins. Biochim Biophys Acta 1212:259–277
Hampton RY (1998) Genetic analysis of hydroxymethylglutaryl-coenzyme A reductase regulated degradation. Curr Opin Lipidol 9:93–97
Hampton RY (2002) ER-associated degradation in protein quality control and cellular regulation. Curr Opin Cell Biol 14:476–482
Hampton RY, Dimster-Denk D, Rine J (1996) The biology of HMG-CoA reductase: the pros of contra-regulation. Trends Biochem Sci 21:140–145
Hastings N, Agaba M, Tocher DR, Leaver MJ, Dick JR, Sargent JR, Teale AJ (2001) A vertebrate fatty acid desaturase with Δ5 and Δ6 activities. Proc Natl Acad Sci USA 98:14304–14309
Heiderpriem RW, Livant PD, Parish EJ, Barbuch RJ, Broaddus MG, Bard M (1992) A simple method for the isolation of zymosterol from a sterol mutant of Saccharomyces cerevisiae. J Steroid Biochem Mol Biol 43:741–743
Holland HL (1999) Recent advances in applied and mechanistic aspects of the enzymatic hydroxylation of steroids by whole-cell biocatalysts. Steroids 64:178–186
Huang YS, Chaudhary S, Thurmond JM, Bobik EG Jr, Yuan L, Chan GM, Kirchner SJ, Mukerji P, Knutzon DS (1999) Cloning of Δ12- and Δ6-desaturases from Mortierella alpina and recombinant production of gamma-linolenic acid in Saccharomyces cerevisiae. Lipids 34:649–659
Hur E, Wilson DK (2000) Crystallization and aldo-keto reductase activity of Gcy1p from Saccharomyces cerevisiae. Acta Crystallogr D Biol Crystallogr 56:763–765
Husselstein T, Gachotte D, Desprez T, Bard M, Benveniste P (1996) Transformation of Saccharomyces cerevisiae with a cDNA encoding a sterol C-methyltransferase from Arabidopsis thaliana results in the synthesis of 24-ethyl sterols. FEBS Lett 381:87–92
Innis SM, Sprecher H, Hachey D, Edmond J, Anderson RE (1999) Neonatal polyunsaturated fatty acid metabolism. Lipids 34:139–149
Jackson BE, Hart-Wells EA, Matsuda SP (2003) Metabolic engineering to produce sesquiterpenes in yeast. Org Lett 5:1629–1632
Jang H, Lin YY, Yang SS (2000) Polyunsaturated fatty acid production with Mortierella alpina by solid substrate fermentation. Bot Bull Acad Sin 41:41–48
Jensen-Pergakes K, Guo Z, Giattina M, Sturley SL, Bard M (2001) Transcriptional regulation of the two sterol esterification genes in the yeast Saccharomyces cerevisiae. J Bacteriol 183:4950–4957
Kajiwara S, Shirai A, Fujii T, Toguri T, Nakamura K, Ohtaguchi K (1996) Polyunsaturated fatty acid biosynthesis in Saccharomyces cerevisiae: expression of ethanol tolerance and the FAD2 gene from Arabidopsis thaliana. Appl Environ Microbiol 62:4309–4313
Kalb VF, Loper JC, Dey CR, Woods CW, Sutter TR (1986) Isolation of a cytochrome P-450 structural gene from Saccharomyces cerevisiae. Gene 45:237–245
Kalb VF, Woods CW, Turi TG, Dey CR, Sutter TR, Loper JC (1987) Primary structure of the P450 lanosterol demethylase gene from Saccharomyces cerevisiae. DNA 6:529–537
Lacour T, Achstetter T, Dumas B (1998) Characterization of recombinant adrenodoxin reductase homologue (Arh1p) from yeast. Implication in in vitro cytochrome p45011beta monooxygenase system. J Biol Chem 273:23984–23992
Lee PC, Schmidt-Dannert C (2002) Metabolic engineering towards biotechnological production of carotinoids in microorganisms. Appl Microbiol Biotechnol 60:1–11
Lees ND, Bard M, Kirsch DR (1999) Biochemistry and molecular biology of sterol synthesis in Saccharomyces cerevisiae. Crit Rev Biochem Mol Biol 34:33–47
Leonard AE, Kelder B, Bobik EG, Chuang LT, Lewis CJ, Kopchick JJ, Mukerji P Huang YS (2002) Identification and expression of mammalian long-chain PUFA elongation enzymes. Lipids 37:733–740
Manzella L, Barros MH, Nobrega FG (1998) ARH1 of Saccharomyces cerevisiae: a new essential gene that codes for a protein homologous to the human adrenodoxin reductase. Yeast 14:839–846
Maraz A (2002) From yeast genetics to biotechnology. Acta Microbiol Immunol Hung 49:483–491
Martin CE, Oh CS, Kandasamy P, Chellapa R, Vemula M (2002) Yeast desaturases. Biochem Soc Trans 30:1080–1082
Meganathan R (2001) Ubiquinone biosynthesis in microorganisms. FEMS Microbiol Lett 203:131–139
Migliaccio A, Castoria G, Di Domenico M, Falco A de, Bilancio A, Lombardi M, Bottero D, Varricchio L, Nanayakkara M, Rotondi A, Auricchio F (2002) Sex steroid hormones act as growth factors. J Steroid Biochem Mol Biol 83:31–35
Miller WL (1995) Mitochondrial specificity of the early steps in steroidogenesis. J Steroid Biochem Mol Biol 55:607–616
Misawa N, Shimada H (1997) Metabolic engineering for the production of carotenoids in non-carotenogenic bacteria and yeasts. J Biotechnol 59:169–181
Morohashi K, Fujii-Kuriyama Y, Okada Y, Sogawa K, Hirose T, Inayama S, Omura T (1984) Molecular cloning and nucleotide sequence of cDNA for mRNA of mitochondrial cytochrome P-450 (SCC) of bovine adrenal cortex. Proc Natl Acad Sci USA 81:4647–4651
Nagasawa N, Bogaki T, Iwamatsu A, Hamachi M, Kumagai C (1998) Cloning and nucleotide sequence of the alcohol acetyltransferase II gene (ATF2) from Saccharomyces cerevisiae Kyokai No. 7. Biosci Biotechnol Biochem 62:1852–1857
Nes WD, McCourt BS, Marshall JA, Ma J, Dennis AL, Lopez M, Li H, He L (1999) Site-directed mutagenesis of the sterol methyl transferase active site from Saccharomyces cerevisiae results in formation of novel 24-ethyl sterols. J Org Chem 64:1535–1542
Novotny C, Beran K, Behalova B, Dolezalova L, Zajicek J (1987) Effect of ammonium ions on Δ5,7-sterol synthesis in Saccharomyces cerevisiae. Folia Microbiol (Praha) 32:206–210
Obeid LM, Okamoto Y, Mao C (2002) Yeast sphingolipids: metabolism and biology. Biochim Biophys Acta 1585:163–171
Oh CS, Toke DA, Mandala S, Martin CE (1997) ELO2 and ELO3, homologues of the Saccharomyces cerevisiae ELO1 gene, function in fatty acid elongation and are required for sphingolipid formation. J Biol Chem 272:17376–17384
Park WS, Murphy PA, Glatz BA (1990) Lipid metabolism and cell composition of the oleaginous yeast Apiotrichum curvatum grown at different carbon to nitrogen ratios. Can J Microbiol 36:318–326
Parker-Barnes JM, Das T, Bobik E, Leonard AE, Thurmond JM, Chaung LT, Huang YS, Mukerji P (2000) Identification and characterization of an enzyme involved in the elongation of n-6 and n-3 polyunsaturated fatty acids. Proc Natl Acad Sci USA. 97:8284–8289
Parks LW, Casey WM (1995) Physiological implications of sterol biosynthesis in yeast. Annu Rev Microbiol 49:95–116
Parks LW, Smith SJ, Crowley JH (1995) Biochemical and physiological effects of sterol alterations in yeast—a review. Lipids 30:227–230
Peyou-Ndi MM, Watts JL, Browse J (2000) Identification and characterization of an animal delta(12) fatty acid desaturase gene by heterologous expression in Saccharomyces cerevisiae. Arch Biochem Biophys 376:399–408
Pirtle IL, Kongcharoensuntorn W, Nampaisansuk M, Knesek JE, Chapman KD, Pirtle RM (2001) Molecular cloning and functional expression of the gene for a cotton Δ12 fatty acid desaturase (FAD2). Biochim Biophys Acta 1522:122–129
Polakowski T, Stahl U, Lang C (1998) Overexpression of a cytosolic hydroxymethylglutaryl-CoA reductase leads to squalene accumulation in yeast. Appl Microbiol Biotechnol 49:66–71
Polakowski T, Bastl R, Stahl U, Lang C (1999) Enhanced sterol-acyl transferase activity promotes sterol accumulation in Saccharomyces cerevisiae. Appl Microbiol Biotechnol 53:30–35
Qiu X, Hong H, MacKenzie SL (2001) Identification of a delta 4 fatty acid desaturase from Thraustochytrium sp. involved in the biosynthesis of docosahexanoic acid by heterologous expression in Saccharomyces cerevisiae and Brassica juncea. J Biol Chem 276:31561–31566
Riad M, Mogos M, Thangathurai D, Lumb PD (2002) Steroids. Curr Opin Crit Care 8:281–284
Rogers B, Decottignies A, Kolaczkowski M, Carvajal E, Balzi E, Goffeau A (2001) The pleiotropic drug ABC transporters from Saccharomyces cerevisiae. J Mol Microbiol Biotechnol 3:207–214
Saito T, Ochiai H (1999) Identification of Delta5-fatty acid desaturase from the cellular slime mold Dictyostelium discoideum. Eur J Biochem 265:809–814
Saito T, Morio T, Ochiai H (2000) A second functional delta5 fatty acid desaturase in the cellular slime mould Dictyostelium discoideum. Eur J Biochem 267:1813–1818
Sakaki T, Kominami S, Hayashi K, Akiyoshi-Shibata M, Yabusaki Y (1996) Molecular engineering study on electron transfer from NADPH-P450 reductase to rat mitochondrial P450c27 in yeast microsomes. J Biol Chem 271:26209–26213
Sakuradani E, Kobayashi M, Ashikari T, Shimizu S (1999) Identification of Δ12-fatty acid desaturase from arachidonic acid-producing Mortierella fungus by heterologous expression in the yeast Saccharomyces cerevisiae and the fungus Aspergillus oryzae. Eur J Biochem 261:812–820
Schenk P, Ausborn M, Bendas F, Nuhn P, Arndt D, Meyer HW (1989) The preparation and characterization of lipid vesicles containing esters of sucrose and fatty acids. J Microencapsul 6:95–103
Sjostrom B, Bergenstahl B, Kronberg B (1993) A method for the preparation of submicron particles of sparingly water-soluble drugs by precipitation in oil-in-water emulsions. II: Influence of the emulsifier, the solvent, and the drug substance. J Pharm Sci 82:584–589
Smith SJ, Crowley JH, Parks LW (1996) Transcriptional regulation by ergosterol in the yeast Saccharomyces cerevisiae. Mol Cell Biol 16:5427–5432
Sorger D, Daum G (2003) Triacylglycerol biosynthesis in yeast. Appl Microbiol Biotechnol 61:289–299
Sprecher H (2000) Metabolism of highly unsaturated n-3 and n-6 fatty acids. Biochim Biophys Acta 1486:219–231
Stach D, Zheng YF, Perez AL, Oehlschlager AC, Abe I, Prestwich GD, Hartman PG (1997) Synthesis and inhibition studies of sulfur-substituted squalene oxide analogues as mechanism-based inhibitors of 2,3-oxidosqualene-lanosterol cyclase. J Med Chem 40:201–209
Sturley SL (2000) Conservation of eukaryotic sterol homeostasis: new insights from studies in budding yeast. Biochim Biophys Acta 1529:155–163
Subbiah MT, Abplanalp W (2003) Ergosterol (major sterol of baker′s and brewer′s yeast extracts) inhibits the growth of human breast cancer cells in vitro and the potential role of its oxidation products. Int J Vitam Nutr Res 73:19–23
Suga K, Honjoh K, Furuya N, Shimizu H, Nishi K, Shinohara F, Hirabaru Y, Maruyama I, Miyamoto T, Hatano S, Iio M (2002) Two low-temperature-inducible Chlorella genes for delta12 and omega-3 fatty acid desaturase (FAD): isolation of delta12 and omega-3 fad cDNA clones, expression of delta12 fad in Saccharomyces cerevisiae, and expression of omega-3 fad in Nicotiana tabacum. Biosci Biotechnol Biochem 66:1314–1327
Swain E, Baudry K, Stukey J, McDonough V, Germann M, Nickels JT Jr (2002) Sterol-dependent regulation of sphingolipid metabolism in Saccharomyces cerevisiae. J Biol Chem 277:26177–26184
Szczebara FM, Chandelier C, Villeret C, Masurel A, Bourot S, Duport C, Blanchard S, Groisillier A, Testet E, Costaglioli P, Cauet G, Degryse E, Balbuena D, Winter J, Achstetter T, Spagnoli R, Pompon D, Dumas B (2003) Total biosynthesis of hydrocortisone from a simple carbon source in yeast. Nat Biotechnol 21:143–149
Thompson BT (2003) Glucocorticoids and acute lung injury. Crit Care Med 31:253–257
Thorsness M, Schafer W, D′Ari L, Rine J (1989) Positive and negative transcriptional control by heme of genes encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase in Saccharomyces cerevisiae. Mol Cell Biol 9:5702–5712
Tijet N, Helvig C, Pinot F, Le Bouquin R, Lesot A, Durst F, Salaun JP, Benveniste I (1998) Functional expression in yeast and characterization of a clofibrate-inducible plant cytochrome P-450 (CYP94A1) involved in cutin monomers synthesis. Biochem J 332:583–589
Valachovic M, Klobucnikova V, Griac P, Hapala I (2002) Heme-regulated expression of two yeast acyl-CoA:sterol acyltransferases is involved in the specific response of sterol esterification to anaerobiosis. FEMS Microbiol Lett 206:121–125
Vandamme EJ (1992) Production of vitamins, coenzymes and related biochemicals by biotechnological processes. J Chem Technol Biotechnol 53:313–327
Veen M, Stahl U, Lang C (2003) Combined overexpression of genes of the ergosterol biosynthetic pathway leads to accumulation of sterols in Saccharomyces cerevisiae. FEMS Yeast Res 4:87–95
Vinson GP, Teja R, Ho MM, Puddefoot JR (1995) A two cell type theory for aldosterone biosynthesis: the roles of 11 beta-hydroxylase and aldosterone synthase, and a high capacity tightly binding steroid carrier for 18-hydroxydeoxycorticosterone in rat adrenals. J Endocrinol 144:359–368
Volkman JK (2003) Sterols in microorganisms. Appl Microbiol Biotechnol 60:495–506
Von Schacky C, Dyerberg J (2001) Omega 3 fatty acids. From eskimos to clinical cardiology—what took us so long? World Rev Nutr Diet 88:90–99
Watkins BA, Li Y, Seifert MF (2001) Nutraceutical fatty acids as biochemical and molecular modulators of skeletal biology. J Am Coll Nutr 20:417–420
Watts JL, Browse J (1999) Isolation and characterization of a Delta 5-fatty acid desaturase from Caenorhabditis elegans. Arch Biochem Biophys 362:175–182
Weber JM, Reiser J, Kappeli O (1990) Lanosterol 14 alpha-demethylase-encoding gene: systematic analysis of homologous overexpression in Saccharomyces cerevisiae using strong yeast promoters. Gene 87:167–175
Weber JM, Ponti CG, Kappeli O, Reiser J (1992) Factors affecting homologous overexpression of the Saccharomyces cerevisiae lanosterol 14 alpha-demethylase gene. Yeast 8:519–533
Woodward RB, Sondhermer F, Taule D, Hensler K, McLamore WH (1952) The total synthesis of steroids. J Am Chem Soc 74:4223
Xu R, Wilson WK, Matsuda SP (2002) Production of meiosis-activating sterols from metabolically engineered yeast. J Am Chem Soc 124:918–919
Yang H, Bard M, Bruner DA, Gleeson A, Deckelbaum RJ, Aljinovic G, Pohl TM, Rothstein R, Sturley SL (1996) Sterol esterification in yeast: a two-gene process Science 272:1353–1356
Zank TK, Zahringer U, Lerchl J, Heinz E (2000) Cloning and functional expression of the first plant fatty acid elongase specific for delta(6)-polyunsaturated fatty acids. Biochem Soc Trans 28:654–658
Zhang B, He X, Tie C, Liu Y (1999) Construction of high ergosterol-producing yeast strains and study on the optimal conditions for culture. Chin J Biotechnol 15:43–49
Zuber MX, Mason JI, Simpson ER, Waterman MR (1988) Simultaneous transfection of COS-1 cells with mitochondrial and microsomal steroid hydroxylases: incorporation of a steroidogenic pathway into nonsteroidogenic cells. Proc Natl Acad Sci USA 85:699–703
Zweytick D, Leitner E, Kohlwein SD, Yu C, Rothblatt J, Daum G (2000) Contribution of Are1p and Are2p to steryl ester synthesis in the yeast Saccharomyces cerevisiae. Eur J Biochem 267:1075–1082
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Veen, M., Lang, C. Production of lipid compounds in the yeast Saccharomyces cerevisiae . Appl Microbiol Biotechnol 63, 635–646 (2004). https://doi.org/10.1007/s00253-003-1456-7
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DOI: https://doi.org/10.1007/s00253-003-1456-7