Skip to main content
SpringerLink
Log in

Cytochrome P450 Expression in Yarrowia lipolytica and Its Use in Steroid Biotransformation

  • Chapter
  • First Online:
Yarrowia lipolytica

Part of the book series: Microbiology Monographs ((MICROMONO,volume 25))

Abstract

This review is a first attempt to systematize data on the potential of transgenic Y. lipolytica to catalyze diverse reactions of steroid transformation. The yeast Y. lipolytica was tested as host for P450-catalyzed biotransformation of steroids, including the mammalian P450scc system (three components) and/or the two-component P450c17 system, being functionally active with yeast NADPH-P450 reductase (CPR). New strategies for the construction of recombinant Y. lipolytica strains containing several expression cassettes containing heterologous cDNA (up to 4, in five vectors) under control of the isocitrate lyase (ICL1) promoter have been developed. Characteristics of recombinant Y. lipolytica strains functionally expressing the P450scc system and/or P450c17, being functionally active with yeast NADPH-P450 reductase (YlCPR), are presented.

Functional expression of P450 systems in yeasts was proved by biotransformation of cholesterol (Cho) or by 17α-hydroxylation of progesterone (Pro) or pregnenolone (Pre). Strains coexpressing the P450scc system and P450c17 exhibited a high biotransformation capacity of Pro into 17α-hydroxyprogesterone (17HPro); the conversion of Cho to Pre and 17α-hydroxypregnenolone occurred rather slowly. For selected P450c17 expressing Y. lipolytica strains, the cultivation conditions (induction, bioconversion) were optimized for high product yield (up to 95 % 17HPro) and reduction of the diol side-product formation from 19–22 % to 1–2 % without gene destruction.

The results obtained could be used for elaboration of new biotechnological approaches with using recombinant yeast strains for synthesis of pharmaceutically active steroids and for screening of compounds which inhibit the P450c17 enzyme activity, playing important roles in the development of hormonal carcinogenesis.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Abbreviations

3β-HSD:

3β-hydroxysteroid dehydrogenase/Δ5,4 isomerase (3β-hydroxy-5-ene steroid dehydrogenase, EC 1.1.1.145)

AdR:

FAD-containing NADPH-adrenodoxin reductase (EC 1.18.1.2)

Adx or shortly Ax:

Adrenodoxin, [2Fe-2S]-ferredoxin from adrenal cortex

AxP:

Fusion of human Adx and P450scc

C. :

Candida

CPR:

NADPH-cytochrome P450-reductase

DHEA:

Dehydroepiandrosterone

E. :

Escherichia

HSD:

Hydroxysteroid dehydrogenase

P450 or CYP:

Cytochrome P450

P450c11 (CYP11B1, P45011β):

Cytochrome P450 11β-hydroxylase

P450c17 (CYP17A1):

Shortly P17 (CYP17, P45017α), cytochrome P450 17α-hydroxylase/17,20-lyase (EC 1.14.99.9)

P450c18 (CYP11B2):

Cytochrome P450 aldosterone synthase

P450c19 (CYP19, P450arom):

Cytochrome P450 aromatase

P450c21 (CYP21):

Cytochrome P450 21-hydroxylase

P450scc (CYP11A1):

Shortly Pb (bovine) or Ph (human), cytochrome P450 cholesterol hydroxylase/20,22-lyase (EC 1.14.15.6)

S. :

Saccharomyces

Y.:

Yarrowia

References

  • Agalarov JA (2008) Coexpression of the cholesterol hydroxylase/lyase system components in Yarrowia lipolytica cells. Diploma thesis, Biochemistry Department, Biological Faculty, Lomonosov Moscow State University

    Google Scholar 

  • Ahmad S, Garg SK, Johri BN (1992) Biotransformation of sterols: selective cleavage of the side chain. Biotechnol Adv 10:1–67

    PubMed  CAS  Google Scholar 

  • Andersen MD, Møller BL (2002) Use of methylotrophic yeast Pichia pastoris for expression of cytochromes P450. Methods Enzymol 357:333–342

    PubMed  CAS  Google Scholar 

  • Andersen MD, Busk PK, Svendsen I, Møller BL (2000) Cytochromes P-450 from cassava (Manihot esculenta Crantz) catalyzing the first steps in the biosynthesis of the cyanogenic glucosides linamarin and lotaustralin. Cloning, functional expression in Pichia pastoris, and substrate specificity of the isolated recombinant enzymes. J Biol Chem 275:1966–1975

    PubMed  CAS  Google Scholar 

  • Athenstaedt K, Jolivet P, Boulard C, Zivy M, Negroni L, Nicaud J-M, Chardot T (2006) Lipid particle composition of the yeast Yarrowia lipolytica depends on the carbon source. Proteomics 6:1450–1459

    PubMed  CAS  Google Scholar 

  • Bankar AV, Kumar AR, Zinjarde SS (2009) Environmental and industrial application of Yarrowia lipolytica. Minireview. Appl Microbiol Biotechnol 84:847–865

    PubMed  CAS  Google Scholar 

  • Barros MH, Nobrega FG (1999) YAH1 of Saccharomyces cerevisiae: a new essential gene that codes for a protein homologous to human adrenodoxin. Gene 233:197–203

    PubMed  CAS  Google Scholar 

  • Barth G, Gaillardin C (1996) Yarrowia lipolytica. In: Wolf K (ed) Nonconventional yeasts in biotechnology. Springer, Heidelberg, pp 313–388

    Google Scholar 

  • 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

    PubMed  CAS  Google Scholar 

  • Beopoulos A, Nicaud JM, Gaillardin C (2011) An overview of lipid metabolism in yeasts and its impact on biotechnological processes. Appl Microbiol Biotechnol 90:1193–1206

    PubMed  CAS  Google Scholar 

  • Bernhardt R (2006) Cytochrome P450 as versatile biocatalysts. J Biotechnol 124:128–145

    PubMed  CAS  Google Scholar 

  • Bourel G, Nicaud J-M, Nthangeni B, Santiago-Gomez P, Belin J-M, Husson F (2004) Fatty acid hydroperoxide lyase of green bell pepper: cloning in Yarrowia lipolytica and biogenesis of volatile aldehydes. Enzyme Microb Technol 35:293–299

    CAS  Google Scholar 

  • Boyle SM, Popp MP, Smith WC, Greenberg RM, James MO (1998) Expression of CYP2L1 in the yeast Pichia pastoris, and determination of catalytic activity with progesterone and testosterone. Mar Environ Res 46:25–28

    CAS  Google Scholar 

  • Braun A, Geier M, Buehler B, Schmid A, Mauersberger S, Glieder A (2012) Steroid biotransformation in biphasic systems with Yarrowia lipolytica expressing human liver cytochrome P450 genes. Microb Cell Fact 11:106. doi:10.1186/1475-2859-11-106

    PubMed  CAS  Google Scholar 

  • Brocard-Masson C, Dumas B (2006) The fascinating world of steroids: S. cerevisiae as a model organism for the study of hydrocortisone biosynthesis. Biotechnol Genet Eng Rev 22:213–252

    PubMed  CAS  Google Scholar 

  • Bureik M, Schiffler B, Hiraoka Y, Vogel F, Bernhardt R (2002) Functional expression of human mitochondrial CYP11B2 in fission yeast and identification of a new internal electron transfer protein, etp1. Biochemistry 41:2311–2321

    PubMed  CAS  Google Scholar 

  • Bureik M, Hübel K, Drăgan CA, Scher J, Becker H, Lenz N, Bernhardt R (2004) Development of test systems for the discovery of selective human aldosterone synthase (CYP11B2) and 11ß-hydroxylase (CYP11B1) inhibitors. Discovery of a new lead compound for the therapy of congestive heart failure, myocardial fibrosis and hypertension. Mol Cell Endocrinol 217:249–254

    PubMed  CAS  Google Scholar 

  • 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

    PubMed  CAS  Google Scholar 

  • Chang Q, Griest TA, Harter TM, Petrash JM (2007) Functional studies of aldo-keto reductases in Saccharomyces cerevisiae. Biochim Biophys Acta 1773:321–329

    PubMed  CAS  Google Scholar 

  • Coelho MAZ, Amaral PFF, Belo I (2010) Yarrowia lipolytica: an industrial workhorse. In: Méndez-Vilas A (ed) Current research, technology and education topics in applied microbiology and microbial biotechnology, vol 2. Formatex, Badajoz, pp 930–944

    Google Scholar 

  • Cornelissen S, Julsing MK, Schmid A, Bühler B (2012) Comparison of microbial hosts and expression systems for mammalian CYP1A1 catalysis. J Ind Microbiol Biotechnol 39:275–287

    PubMed  CAS  Google Scholar 

  • Crowley JH, Leak FW, Shianna KV, Tove S, Parks LW (1998) A mutation in a purported regulatory gene affects control of sterol uptake in Saccharomyces cerevisiae. J Bacteriol 180:4177–4183

    PubMed  CAS  Google Scholar 

  • Czabany T, Athenstaedt K, Daum G (2007) Synthesis, storage and degradation of neutral lipids in yeast. Biochim Biophys Acta 1771:299–309

    PubMed  CAS  Google Scholar 

  • Dietrich M, Grundmann L, Kurr K, Valinotto L, Saussele T, Schmid RD, Lange S (2005) Recombinant production of human microsomal cytochrome P450 2D6 in the methylotrophic yeast Pichia pastoris. Chembiochem 6:2014–2022

    PubMed  CAS  Google Scholar 

  • Drăgan CA, Zearo S, Hannemann F, Bernhardt R, Bureik M (2005) Efficient conversion of 11-deoxycortisol to cortisol (hydrocortisone) by recombinant fission yeast Schizosaccharomyces pombe. FEMS Yeast Res 5:621–625

    PubMed  Google Scholar 

  • Drăgan CA, Hartmann RW, Bureik M (2006) A fission yeast-based test system for the determination of IC50 values of anti-prostate tumor drugs acting on CYP21. J Enzyme Inhib Med Chem 21:547–556

    PubMed  Google Scholar 

  • Dujon B, Sherman D, Fischer G et al (2004) Genome evolution in yeasts. Nature 430:35–44

    PubMed  Google Scholar 

  • Dumas B, Cauet G, Lacour T, Degryse E, Laruelle L, Ledoux C, Spagnoli R, Achstetter T (1996) 11 beta-hydroxylase activity in recombinant yeast mitochondria. In vivo conversion of 11-deoxycortisol to hydrocortisone. Eur J Biochem 238:495–504

    PubMed  CAS  Google Scholar 

  • Dumas B, Brocard-Masson C, Assemat-Lebrun K, Achstetter T (2006) Hydrocortisone made in yeast: metabolic engineering turns a unicellular microorganism into a drug-synthesizing factory. Biotechnol J 1:299–307

    PubMed  CAS  Google Scholar 

  • Duport C, Spagnoli R, Degryse E, Pompon D (1998) Self-sufficient biosynthesis of pregnenolone and progesterone in engineered yeast. Nat Biotechnol 16:186–189

    PubMed  CAS  Google Scholar 

  • 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

    PubMed  CAS  Google Scholar 

  • Dutta TK, Datta J, Samanta TB (1993) Onset of new catalytic activity in immobilized spores of Aspergillus ochraceus TS due to in situ germination: C17-C20 lysis accompanies 11 alpha-hydroxylation of steroid. Biochem Biophys Res Commun 192:119–123

    PubMed  CAS  Google Scholar 

  • Ehmer PB, Bureik M, Bernhardt R, Müller U, Hartmann RW (2002) Development of a test system for inhibitors of human aldosterone synthase (CYP11B2): screening in fission yeast and evaluation of selectivity in V79 cells. J Steroid Biochem Mol Biol 81:173–179

    PubMed  CAS  Google Scholar 

  • Ewen KM, Schiffler B, Uhlmann-Schiffler H, Bernhardt R, Hannemann F (2008) The endogenous adrenodoxin reductase-like flavoprotein arh1 supports heterologous cytochrome P450-dependent substrate conversions in Schizosaccharomyces pombe. FEMS Yeast Res 8:432–441

    PubMed  CAS  Google Scholar 

  • Faletrov YV, Frolova NS, Rudaya EV, Radyuk VG, Shkumatov VM (2008a) Inhibition of pregnenolone and related steroids 3β-acetylation in recombinant yeasts Saccharomyces cerevisiae GRF18/YEp5117α by aliphatic alcohols. In: Kilchevskii AV (ed) Molecular and applied genetics: Proceedings, vol 7, Minsk, pp 45–48

    Google Scholar 

  • Faletrov YV, Frolenkov KA, Novikova LA, Luzikov VN, Mauersberger S, Barth G, Shkumatov VM (2008b) Transgenic yeasts for pharmacological investigations. In: 9th International symposium on cytochrome P450 biodiversity and biotechnology, Nice, France, 08–12 June 2008, Abstracts

    Google Scholar 

  • Fallon RD, Payne MS, Picataggio SK, Wu S (1999) Transformed yeast strains and their use for the production of monoterminal and diterminal aliphatic carboxylates. WO9904014

    Google Scholar 

  • Fallon RD, Payne MS, Picataggio SK, Wu S (2004) Transformed yeast strains and their use for the production of monoterminal and diterminal aliphatic carboxylates. US2004146999

    Google Scholar 

  • Fernandes P, Cruz A, Angelova B, Pinheiro HM, Cabral JMS (2003) Microbial conversion of steroid compounds: recent developments. Enzyme Microb Technol 32:688–705

    CAS  Google Scholar 

  • Fickers P, Benetti PH, Wache Y, Marty A, Mauersberger S, Smit MS, Nicaud JM (2005) Hydrophobic substrate utilisation by the yeast Yarrowia lipolytica, and its potential applications. FEMS Yeast Res 5:527–543

    PubMed  CAS  Google Scholar 

  • Förster A (2001) Untersuchungen zur heterologen Expression von steroidwandelnden Cytochrom P450 Systemen in der Hefe Yarrowia lipolytica. Diploma thesis, Technische Universität Dresden, Germany

    Google Scholar 

  • Fukuda R, Ohta A (2013) Utilization of hydrophobic substrates by Yarrowia lipolytica. In: Barth G (ed) Yarrowia lipolytica, vol 1. Springer, Heidelberg

    Google Scholar 

  • Gerber J (1999) Untersuchungen zur Optimierung des Elektronentransportsystems für die Cytochrom P450 katalysierte Biotransformation von Steroiden in Yarrowia lipolytica. Diploma thesis, Technische Universität Dresden

    Google Scholar 

  • Gonzalez FJ, Korzekwa KR (1995) Cytochromes P450 expression systems. Annu Rev Pharmacol Toxicol 35:369–390

    PubMed  CAS  Google Scholar 

  • Hakki T, Zearo S, Drăgan CA, Bureik M, Bernhardt R (2008) Coexpression of redox partners increases the hydrocortisone (cortisol) production efficiency in CYP11B1 expressing fission yeast Schizosaccharomyces pombe. J Biotechnol 133:351–359

    PubMed  CAS  Google Scholar 

  • Juretzek T (1999) Entwicklung von Wirts-Vektor-Systemen zur heterologen Expression von Proteinen in der nichtkonventionellen Hefe Yarrowia lipolytica und ihre Anwendung für die Cytochrom P450-katalysierte Stoffumwandlung. PhD thesis, Technische Universität Dresden

    Google Scholar 

  • Juretzek T, Prinz A, Schunck W-H, Barth G, Mauersberger S (1997) Expressionskassetten zur heterologen Expression von Proteinen in der Hefe Yarrowia lipolytica unter Kontrolle des regulierbaren Promotors der Isocitratlyase. Patent DE19525282

    Google Scholar 

  • Juretzek T, Mauersberger S, Schunck W-H, Nicaud J-M, Barth G (1999) Functional expression of heterologous cytochrome P450 in the yeast Yarrowia lipolytica and its use in biotransformation of steroids. Curr Genet 35(3–4):307

    Google Scholar 

  • Juretzek T, Wang H-J, Nicaud J-M, Mauersberger S, Barth G (2000a) Comparison of promoters suitable for regulated overexpression of heterologous genes in the alkane-utilizing yeast Yarrowia lipolytica. Biotechnol Bioprocess Eng 5:320–326

    CAS  Google Scholar 

  • Juretzek T, Mauersberger S, Barth G (2000b) Recombinant haploid or diploid Yarrowia lipolytica cells for the functional heterologous expression of cytochrome P450 systems. Patent WO0003008, DE19932811

    Google Scholar 

  • Juretzek T, Le Dall MT, Mauersberger S, Gaillardin C, Barth G, Nicaud J-M (2001) Vectors for gene expression and amplification in the yeast Yarrowia lipolytica. Yeast 18:97–113

    PubMed  CAS  Google Scholar 

  • Katsumata T, Hasegawa A, Fujiwara T, Komatsu T, Notomi M, Abe H, Natsume M, Kawaide H (2008) Arabidopsis CYP85A2 catalyzes lactonization reactions in the biosynthesis of 2-deoxy-7-oxalactone brassinosteroids. Biosci Biotechnol Biochem 72:2110–2117

    PubMed  CAS  Google Scholar 

  • Ko KW, Lin F, Katsumata T, Sugai Y, Miyazaki S, Kawaide H, Okada K, Nojiri H, Yamane H (2008) Functional identification of a rice ent-kaurene oxidase, OsKO2, using the Pichia pastoris expression system. Biosci Biotechnol Biochem 72:3285–3288

    PubMed  CAS  Google Scholar 

  • Kolar NW, Swart AC, Mason JI, Swart P (2007) Functional expression and characterisation of human cytochrome P45017α in Pichia pastoris. J Biotechnol 129:635–644

    PubMed  CAS  Google Scholar 

  • Kovganko NV, Kashkan ZN, Shkumatov VM (2001) Simple synthesis of 17α,20β-dihydroxypregn-4-en-3-one. Chem Nat Comp 37:55–56

    CAS  Google Scholar 

  • Lieberman S, Warne PA (2001) 17-Hydroxylase: an evaluation of the present view of its catalytic role in steroidogenesis. J Steroid Biochem Mol Biol 78:299–312

    PubMed  CAS  Google Scholar 

  • Lu W, Du L, Wang M, Guo Y, Lu F, Sun B, Wen J, Jia X (2007) A novel substrate addition method in the 11ß-hydroxylation of steroids by Curvularia lunata. Food Bioprod Process 85:63–72

    CAS  Google Scholar 

  • Manosroi J, Saowakhon S, Manosroi A (2008) Enhancement of 17α-hydroxyprogesterone production from progesterone by biotransformation using hydroxypropyl-β-cyclodextrin complexation technique. J Steroid Biochem Mol Biol 112:201–204

    PubMed  CAS  Google Scholar 

  • Masuda Y, Park SM, Ohkuma M, Ohta A, Takagi M (1994) Expression of an endogenous and a heterologous gene in Candida maltosa by using a promoter of a newly-isolated phosphoglycerate kinase (PGK) gene. Curr Genet 25:412–417

    PubMed  CAS  Google Scholar 

  • Matsuzaki F, Wariishi H (2005) Molecular characterization of cytochrome P450 catalyzing hydroxylation of benzoates from the white-rot fungus Phanerochaete chrysosporium. Biochem Biophys Res Commun 334:1184–1190

    PubMed  CAS  Google Scholar 

  • Mauersberger S (2013) Cytochrom P450 of the alkane-utilizing yeast Yarrowia lipolytica. In: Barth G (ed) Yarrowia lipolytica. Springer, Heidelberg

    Google Scholar 

  • Mauersberger S, Kärgel E, Matyashova RN, Müller H-G (1987) Subcellular organization of alkane oxidation in Candida maltosa. J Basic Microbiol 27:565–582

    CAS  Google Scholar 

  • Mauersberger S, Ohkuma M, Schunck W-H, Takagi M (1996) Chapter 12. Candida maltosa. In: Wolf K (ed) Non-conventional yeasts in biotechnology. A handbook. Springer, Berlin, pp 411–580

    Google Scholar 

  • Mauersberger S, Wang H-J, Gaillardin C, Barth G, Nicaud J-M (2001) Insertional mutagenesis in the n-alkane-assimilating yeast Yarrowia lipolytica: generation of tagged mutants in genes involved in hydrophobic substrates utilization. J Bacteriol 183:5102–5109

    PubMed  CAS  Google Scholar 

  • Mauersberger S, Förster A, Gerber J, Juretzek T, Barth G (2002) Functional expression of heterologous cytochrome P450 in Yarrowia lipolytica and its use in biotransformation of steroids. In: Third Yarrowia lipolytica international meeting (TYLIM2002), Dresden, Germany, 17–20 July 2002, Abstracts p 114 (Poster 41)

    Google Scholar 

  • McCormick SP, Alexander NJ, Proctor RH (2006) Heterologous expression of two trichothecene P450 genes in Fusarium verticillioides. Can J Microbiol 52:220–226

    PubMed  CAS  Google Scholar 

  • Menke HH, Nan LM, Selten GCM, Smaal EB, Slijkhuis H (1990) Cloning and co-expression of bovine adrenal cytochrome P450scc and adrenodoxin in Kluyveromyces lactis. In: 20th FEBS-Meeting, Budapest, Hungary, 19–24 August 1990, program p 215, poster P-TH-381

    Google Scholar 

  • Menzel R, Vogel F, Kärgel E, Schunck W-H (1997) Inducible membranes in yeast: relation to the unfolded-protein-response pathway. Yeast 13:1211–1229

    PubMed  CAS  Google Scholar 

  • Minenko AN, Novikova LA, Luzikov VN, Kovaleva IE (2008) Import of hybrid forms of CYP11A1 into yeast mitochondria. Biochim Biophys Acta 1780:1121–1130

    PubMed  CAS  Google Scholar 

  • Ness F, Achstetter T, Duport C, Karst F, Spagnoli R, Degryse E (1998) Sterol uptake in Saccharomyces cerevisiae heme auxotrophic mutants is affected by ergosterol and oleate but not by palmitoleate or by sterol esterification. J Bacteriol 180:1913–1919

    PubMed  CAS  Google Scholar 

  • Novikova LA, Yovkova V, Faletrov YV, Agalarov JA, Luzikov VN, Shkumatov VM, Barth G, Mauersberger S (2008) Functional expression of the cholesterol side-chain cleavage cytochrome P450scc system and of P450c17 in the yeast Yarrowia lipolytica. In: 9th International symposium on cytochrome P450 biodiversity and biotechnology, Nice, France, 08–12 June 2008, Abstracts p 68

    Google Scholar 

  • Novikova LA, Faletrov YV, Kovaleva IE, Mauersberger S, Luzikov VN, Shkumatov VM (2009) From structure and functions of steroidogenic enzymes to new technologies of gene engineering. Review. Biochemistry (Moscow) 74:1482–1504

    CAS  Google Scholar 

  • Nthangeni MB, Urban P, Pompon D, Smit M, Nicaud J-M (2004) The use Yarrowia lipolytica for the expression of human cytochrome P450 CYP1A1. Yeast 21:583–592

    PubMed  CAS  Google Scholar 

  • Oeda K, Sakaki T, Ohkawa H (1985) Expression of rat liver cytochrome P-450MC cDNA in Saccharomyces cerevisiae. DNA 4:203–210

    PubMed  CAS  Google Scholar 

  • Ohkuma M, Kobayashi K, Kawai S, Hwang CW, Ohta A, Takagi M (1995a) Identification of a centromeric activity in the autonomously replicating TRA region allows improvement of the host-vector system of Candida maltosa. Mol Gen Genet 249:447–455

    PubMed  CAS  Google Scholar 

  • Ohkuma M, Park S-M, Zimmer T, Menzel R, Vogel F, Schunck W-H, Ohta A, Takagi M (1995b) Proliferation of intracellular membrane structures upon homologous overproduction of cytochrome P-450 in Candida maltosa. Biochim Biophys Acta (Biomembranes) 1236:163–169

    Google Scholar 

  • Ortiz de Montellano PR (ed) (2005) Cytochrome P450: structure, mechanism and biochemistry, 3rd edn. Plenum, New York

    Google Scholar 

  • Pajic T, Vitas M, Zigon D, Pavko A, Kelly S, Komel R (1999) Biotransformation of steroids by the fission yeast Schizosaccharomyces pombe. Yeast 15:639–645

    PubMed  CAS  Google Scholar 

  • Park SM, Ohkuma M, Masuda Y, Ohta A, Takagi M (1997) Galactose-inducible expression systems in Candida maltosa using promoters of newly-isolated GAL1 and GAL10 genes. Yeast 13:21–29

    PubMed  CAS  Google Scholar 

  • Peters FT, Drăgan CA, Wilde DR, Meyer MR, Zapp J, Bureik M, Maurer HH (2007) Biotechnological synthesis of drug metabolites using human cytochrome P450 2D6 heterologously expressed in fission yeast exemplified for the designer drug metabolite 4′-hydroxymethyl-alpha-pyrrolidinobutyrophenone. Biochem Pharmacol 74:511–520

    PubMed  CAS  Google Scholar 

  • Peters FT, Drăgan CA, Schwaninger AE, Sauer C, Zapp J, Bureik M, Maurer HH (2009a) 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

    PubMed  CAS  Google Scholar 

  • Peters FT, Drăgan CA, Kauffels A, Schwaninger AE, Zapp J, Bureik M, Maurer HH (2009b) Biotechnological synthesis of the designer drug metabolite. 4′-hydroxymethyl-alpha-pyrrolidinohexanophenone in fission yeast heterologously expressing human cytochrome P450 2D6 – a versatile alternative to multistep chemical synthesis. J Anal Toxicol 33:190–197

    PubMed  CAS  Google Scholar 

  • Picataggio S, Rohrer T, Deanda K, Lanning D, Reynolds R, Mielenz J, Eirich LD (1992) Metabolic engineering of Candida tropicalis for the production of long-chain dicarboxylic acids. Biotechnology (NY) 10:894–898

    CAS  Google Scholar 

  • Pikuleva IA (2006) Cytochrome P450s and cholesterol homeostasis. Pharmacol Ther 112:761–773

    PubMed  CAS  Google Scholar 

  • Pompon D, Louerat B, Bronine A, Urban P (1996) Yeast expression of animal and plant P450s in optimized redox environments. Methods Enzymol 272:51–64

    PubMed  CAS  Google Scholar 

  • Pompon D, Gautier JC, Perret A, Truan G, Urban P (1997) Simulation of human xenobiotic metabolism in microorganisms. Yeast a good compromise between E. coli and human cells. Review. J Hepatol 26(Suppl 2):81–85

    PubMed  CAS  Google Scholar 

  • Prinz A (1995) Testung des ICL1-Promoters zur funktionellen heterologen Expression von Cytochrom P450 in der Hefe Yarrowia lipolytica. Universität Gesamthochschule Kassel, Fachbereich 19 (Biologie/Chemie)

    Google Scholar 

  • Raychaudhuri S, Prinz WA (2006) Uptake and trafficking of exogenous sterols in Saccharomyces cerevisiae. Biochem Soc Trans 34:359–362

    PubMed  CAS  Google Scholar 

  • Renaud JP, Peyronneau MA, Urban P, Truan G, Cullin C, Pompon D, Beaune P, Mansuy D (1993) Recombinant yeast in drug metabolism. Review. Toxicology 82:39–52

    PubMed  CAS  Google Scholar 

  • Rudaya EV (2007) Catalytic efficiency and substrate specificity of recombinant yeast expressing cytochrome Р450с17. PhD Thesis, September 2007, Byelorussian State University

    Google Scholar 

  • Rudaya EV, Frolova NS, Mauersberger S, Shkumatov VM (2006) Catalytic efficiency of progesterone biotransformation by recombinant Yarrowia lipolytica yeasts. In: International conference on molecular, membrane and basic cellular principles of functions of biosystems (VII. congress of BOOFiB = BSSPhB), 21–23 June 2006, Minsk, Belarus, Poster I, pp 129–131

    Google Scholar 

  • Sakaki T, Inouye K (2000) Practical application of mammalian cytochrome P450. Review. J Biosci Bioeng 90:583–590

    PubMed  CAS  Google Scholar 

  • Sakaki T, Shibata M, Yabusaki Y, Murakami H, Ohkawa H (1989) Expression of bovine cytochrome P450c17 cDNA in Saccharomyces cerevisiae. DNA 8:409–418

    PubMed  CAS  Google Scholar 

  • Sakaki T, Shibata M, Yabusaki Y, Murakami H, Ohkawa H (1990) Expression of bovine cytochrome P450c21 and its fused enzymes with yeast NADPH-cytochrome P450 reductase in Saccharomyces cerevisiae. DNA Cell Biol 9:603–614

    PubMed  CAS  Google Scholar 

  • Sakaki T, Akiyoshi-Shibata M, Yabusaki Y, Manabe K, Murakami H, Ohkawa H (1991) Progesterone metabolism in recombinant yeast simultaneously expressing bovine cytochromes P450c17 (CYP17A1) and P450c21 (CYP21B1) and yeast NADPH-P450 oxidoreductase. Pharmacogenetics 1:86–93

    PubMed  CAS  Google Scholar 

  • 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 271:2609–26213

    Google Scholar 

  • Sandig G, Kärgel E, Menzel R, Vogel F, Zimmer T, Schunck WH (1999) Regulation of endoplasmic reticulum biogenesis in response to cytochrome P450 overproduction. Drug Metab Rev 31:393–410

    PubMed  CAS  Google Scholar 

  • Santiago-Gómez MP, Vergely C, Policar C, Nicaud J-M, Belin J-M, Rochette L, Husson F (2007) Characterization of purified green bell pepper hydroperoxide lyase expressed by Yarrowia lipolytica: radicals detection during catalysis. Enzyme Microb Technol 41:13–18

    Google Scholar 

  • Schiffler B, Bureik M, Reinle W, Müller EC, Hannemann F, Bernhardt R (2004) The adrenodoxin-like ferredoxin of Schizosaccharomyces pombe mitochondria. J Inorg Biochem 98:1229–1237

    PubMed  CAS  Google Scholar 

  • Schunck W-H, Mauersberger S, Huth J, Riege P, Müller H-G (1987a) Function and regulation of cytochrome P-450 in alkane-assimilating yeast I. Selective inhibition with carbon monoxide in growing cells. Arch Microbiol 147:240–244

    CAS  Google Scholar 

  • Schunck W-H, Mauersberger S, Kärgel E, Huth J, Müller H-G (1987b) Function and regulation of cytochrome P-450 in alkane-assimilating yeast II. Effect of oxygen-limitation. Arch Microbiol 147:245–248

    CAS  Google Scholar 

  • Schunck W-H, Vogel F, Gross B, Kärgel E, Mauersberger S, Köpke K, Gengnagel C, Müller H-G (1991) Comparison of two cytochromes P-450 from Candida maltosa: primary structures, substrate specificities, and effects of their expression in Saccharomyces cerevisiae on the proliferation of the endoplasmic reticulum. Eur J Cell Biol 55:336–345

    PubMed  CAS  Google Scholar 

  • Sedlaczek L (1988) Biotransformation of steroids. Crit Rev Biotechnol 7:187–236

    PubMed  CAS  Google Scholar 

  • Shiningavamwe A, Obiero G, Albertyn J, Nicaud J-M, Smit M (2006) Heterologous expression of the benzoate para-hydroxylase encoding gene (CYP53B1) from Rhodotorula minuta by Yarrowia lipolytica. Appl Microbiol Biotechnol 72:323–329

    PubMed  CAS  Google Scholar 

  • Shkumatov VM, Smettan G (1991) Heterologous reconstruction of monooxygenase systems: “Substrate specific cytochrome P-450-microorganism” (Russian). In: Finogenova TV, Sharyshev AA (eds) Alkane metabolism and oversynthesis of metabolites by microorganisms. Center for Biological Research, USSR Academy of Sciences, Pushchino, Russia, pp 78–86

    Google Scholar 

  • Shkumatov VM, Radyuk VG, Usova EV, Schunck W-H, Mauersberger S (1998) Transgenic strains of microorganisms in the biosynthesis of steroids (Russian). In: Sviridov VV (ed) Chemical problems in the creation of new materials and technologies. Minsk University Press, Minsk, pp 559–566

    Google Scholar 

  • Shkumatov VM, Usova EV, Poljakov YS, Frolova NS, Radyuk VG, Mauersberger S, Chernogolov AA, Honeck H, Schunck W-H (2002) Biotransformation of steroids by a recombinant yeast strain expressing bovine cytochrome P-45017α. Biochemistry (Moscow) 67:456–467

    CAS  Google Scholar 

  • Shkumatov VM, Usova EV, Radyuk VG, Kashkan ZN, Kovganko NV, Juretzek T, Mauersberger S (2003) Oxidation of 17α,20ß- and 17α,20α-dihydroxypregn-4-en-3-ones, side products of progesterone biotransformation with recombinant microorganisms, expressing cytochrome P-45017α. Russ J Bioorgan Chem 29:581–587

    CAS  Google Scholar 

  • Shkumatov VM, Frolova NS, Rudaya EV, Faletrov YV, Mauersberger S, Barth G (2006) Range of substrates and steroid bioconversion reactions performed by recombinant microorganisms Saccharomyces cerevisiae and Yarrowia lipolytica expressing cytochrome P450c17. Appl Biochem Microbiol (Moscow) 42:472–478

    CAS  Google Scholar 

  • Shkumatov VM, Falertov YV, Mauersberger S, Barth G, Novikova LA, Luzikov VN (2007a) Recombinant microorganisms for the synthesis of steroids and testing of medicines. EMBO Workshop The chemistry and biochemistry of catalysis by biological systems, 20–22 June 2007, EMBL Hamburg, Abstracts p 20 (invited oral presentation) and p46

    Google Scholar 

  • Shkumatov VM, Faletrov YV, Mauersberger S, Barth G, Novikova LA, Luzikov VN (2007b) New approaches to the synthesis of steroids and testing of medical compounds. In: XVIIIth Mendeleev congress on general and applied chemistry, 23–28 Sept 2007, Moscow, Russia (invited oral presentation)

    Google Scholar 

  • Shkumatov VM, Usova EV, Frolova NS, Barth G, Mauersberger S (2007c) Effect of steroid biosynthesis modifiers on the progesterone biotransformation by recombinant yeasts expressing cytochrome P450c17. Biochem (Moscow) Suppl Ser B Biomed Chem 1:87–94

    Google Scholar 

  • Slijkhuis H, Selten GCM Smaal EB (1989) Process for the biochemical oxidation of steroids and genetically engineered cells used therefor. Patent EP0340878

    Google Scholar 

  • Slijkhuis H, Smaal EB, Selten GCM (1990–2004) Process for multiple oxidation of steroids and genetically engineered cells used therein. Patent series NL88202080, EP0360361, WO8910963, WO9003428, US20040067579

    Google Scholar 

  • Storbeck KH, Kolar NW, Stander M, Swart AC, Prevoo D, Swart P (2008) The development of an ultra performance liquid chromatography-coupled atmospheric pressure chemical ionization mass spectrometry assay for seven adrenal steroids. Anal Biochem 372:11–20

    PubMed  CAS  Google Scholar 

  • 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

    PubMed  CAS  Google Scholar 

  • Taylor FR, Parks LW (1981) An assessment of the specificity of sterol uptake and esterification in Saccharomyces cerevisiae. J Biol Chem 256:13048–13054

    PubMed  CAS  Google Scholar 

  • Theron CW (2007) Heterologous expression of cytochrome P450 monooxygenases by the yeast Yarrowia lipolytica. M.S. thesis, University of the Free State, Bloemfontein, South Africa

    Google Scholar 

  • Thevenieau (2006) Ingenierie metabolique de la levure Yarrowia lipolytica pour la production d’acides dicarboxyliques a partier d’huiles vegetales. PhD thesis, INRA, Paris-Grignon

    Google Scholar 

  • Thevenieau F, Nicaud J-M, Gaillardin C (2009) Chapter 26. Applications of the non-conventional yeast Yarrowia lipolytica. In: Satyanarayana T, Kunze G (eds) Yeast biotechnology: diversity and applications. Springer, Dordrecht, pp 589–613. doi:10.1007/978-1-4020-8292-4_26

    Google Scholar 

  • Thevenieau F, Beopoulos F, Desfougeres T, Sabirova J, Albertin K, Zinjarde S, Nicaud J-M (2010) Uptake and assimilation of hydrophobic substrates by the oleaginous yeast Yarrowia lipolytica. In: Timmis KN (ed) Handbook of hydrocarbon and lipid microbiology, vol 2, part 7, chap 48. Springer, Berlin, pp 1513–1527

    Google Scholar 

  • Trant JM (1996) Functional expression of recombinant spiny dogfish shark (Squalus acanthias) cytochrome P450c17 (17α-hydroxylase/C17,20-lyase) in yeast (Pichia pastoris). Arch Biochem Biophys 326:8–14

    PubMed  CAS  Google Scholar 

  • Urban P, Truan G, Bellamine A, Laine R, Gautier JC, Pompon D (1994) Engineered yeasts simulating P450-dependent metabolisms: tricks, myths and reality. Review. Drug Metabol Drug Interact 11:169–200

    PubMed  CAS  Google Scholar 

  • Vadali RV, Horton CE, Rudolph FB, Bennett GN, San KY (2004) Production of isoamyl acetate in ackA-pta and/or ldh mutants of Escherichia coli with overexpression of yeast ATF2. Appl Microbiol Biotechnol 63:698–704

    PubMed  CAS  Google Scholar 

  • Vico P, Cauet G, Rose K, Lathe R, Degryse E (2002) Dehydroepiandrosterone (DHEA) metabolism in Saccharomyces cerevisiae expressing mammalian steroid hydroxylase CYP7B: Ayr1p and Fox2p display 17β-hydroxysteroid dehydrogenase activity. Yeast 19:873–886

    PubMed  CAS  Google Scholar 

  • Wilcox LJ, Balderes DA, Wharton B, Tinkelenberg AH, Rao G, Sturley SL (2002) Transcriptional profiling identifies two members of the ATP-binding cassette transporter superfamily required for sterol uptake in yeast. J Biol Chem 277:32466–32472

    PubMed  CAS  Google Scholar 

  • Wilson CR, Craft DL, Eirich LD, Eshoo M, Maduri KK, Cornett CA, Brenner AA, Tang M, Loper JC, Gleeson M (2001) Cytochrome P450 monooxygenase and NADPH cytochrome P450 oxidoreductase genes and proteins related to the omega-hydroxylase complex of Candida tropicalis and methods relating thereto. Patent US6331420

    Google Scholar 

  • Wright R (1993) Insights from inducible membranes. Curr Biol 3:870–873

    PubMed  CAS  Google Scholar 

  • Wu DA, Hu MC, Chung BC (1991) Expression and functional study of wild-type and mutant human cytochrome P450c21 in Saccharomyces cerevisiae. DNA Cell Biol 10:201–209

    PubMed  CAS  Google Scholar 

  • Yamazaki S, Sato K, Suhara K, Sakaguchi M, Mihara K, Omura T (1993) Importance of the proline-rich region following signal-anchor sequence in the formation of correct conformation of microsomal cytochrome P-450s. J Biochem 114:652–657

    PubMed  CAS  Google Scholar 

  • Yasumori T (1997) Expression of a human cytochrome P450 in Schizosaccharomyces pombe: comparison with the expression in Saccharomyces cerevisiae. In: Giga-Hama Y, Kumagai H (eds) Foreign gene expression in fission yeast Schizosaccharomyces pombe. Springer, Berlin, pp 111–121

    Google Scholar 

  • Yasumori T, Chen LS, Li QH, Ueda M, Tsuzuki T, Goldstein JA, Kato R, Yamazoe Y (1999) Human CYP2C-mediated stereoselective phenytoin hydroxylation in Japanese: difference in chiral preference of CYP2C9 and CYP2C19. Biochem Pharmacol 57:1297–1303

    PubMed  CAS  Google Scholar 

  • Yovkova V (2006) Construction of recombinant Yarrowia lipolytica strains for the heterologous expression of the cholesterol-site chain cleaving cytochrome P450 system (German). Diploma Thesis, May 2006, Institute of Microbiology, Dresden University of Technology

    Google Scholar 

  • Yovkova V, Novikova LA, Luzikov VN, Barth G, Mauersberger S (2007) Expression of the cholesterol side-chain cleavage cytochrome P450scc system in Yarrowia lipolytica. In: IV International symposium “intracellular protein catabolism”, VAAM (Association for General and Applied Microbiology) Annual meeting, Osnabrueck, Germany, 1–4 April 2007, Abstracts PB078, pp 103–104, Biospektrum Sonderausgabe Tagungsband 2007

    Google Scholar 

  • Zimmer T, Schunck W-H (1995) A deviation from the universal genetic code in Candida maltosa and consequences for heterologous expression of cytochromes P450 52A4 and 52A5 in Saccharomyces cerevisiae. Yeast 11:33–41

    PubMed  CAS  Google Scholar 

  • Zöllner A, Drăgan CA, Pistorius D, Müller R, Bode HB, Peters FT, Maurer HH, Bureik M (2009) Human CYP4Z1 catalyzes the in-chain hydroxylation of lauric acid and myristic acid. Biol Chem 390:313–317

    PubMed  Google Scholar 

  • Zöllner A, Parr MK, Drăgan CA, Dräs S, Schlörer N, Peters FT, Maurer HH, Schänzer W, Bureik M (2010) CYP21-catalyzed production of the long-term urinary metandienone metabolite 17ß-hydroxymethyl-17α-methyl-18-norandrosta-1,4,13-trien-3-one: a contribution to the fight against doping. Biol Chem 391:119–127

    PubMed  Google Scholar 

Download references

Acknowledgements

Parts of this work were supported by the Russian Foundation for Basic Research (projects 05-04-48049a and 08-04-00842a), the Belorussian Foundation for Basic Research (project BO4MC-030), the Bundesministerium für Bildung und Forschung of Germany (BMBF, former Bundesministerium für Forschung und Technologie, BMFT, projects 0310257 and 0310654), and the INTAS project CA 03-51-4366. The authors would like to thank the PhD and diploma students which have been involved in theses studies (since the middle of the 1990s). We thank Wolf-Hagen Schunck (Berlin) for his support and fruitful discussions during the first steps of this research, and we gratefully acknowledge the permanent interest and support of the late (deceased) Valentin N. Luzikov (Moscow), Jean-Marc Nicaud (Thiverval-Grignon) and of Gerold Barth (Dresden) to this research.

Author information

Authors and Affiliations

  1. Institute of Microbiology, Technische Universität Dresden, 01062, Dresden, Germany

    Stephan Mauersberger

  2. Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119991, Moscow, Russia

    Ludmila A. Novikova

  3. Institute of Physico-Chemical Problems, Belarusian State University, 220030, Minsk, Belarus

    Vladimir M. Shkumatov

  4. Department of Chemistry, Laboratory of Biochemistry of Medical Drugs, 220030, Minsk, Belarus

    Vladimir M. Shkumatov

Authors
  1. Stephan Mauersberger
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ludmila A. Novikova
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Vladimir M. Shkumatov
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Vladimir M. Shkumatov .

Editor information

Editors and Affiliations

  1. TU Dresden Inst. Mikrobiologie, Dresden, Sachsen, Germany

    Gerold Barth

Rights and permissions

Reprints and permissions

Copyright information

© 2013 Springer-Verlag Berlin Heidelberg

About this chapter

Cite this chapter

Mauersberger, S., Novikova, L.A., Shkumatov, V.M. (2013). Cytochrome P450 Expression in Yarrowia lipolytica and Its Use in Steroid Biotransformation. In: Barth, G. (eds) Yarrowia lipolytica. Microbiology Monographs, vol 25. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-38583-4_7

Download citation

Publish with us

Policies and ethics

Search

Navigation