Molecular Genetics and Genomics

, Volume 274, Issue 3, pp 283–294 | Cite as

Characterization of the oat1 gene of Penicillium chrysogenum encoding an ω-aminotransferase: induction by L-lysine, L-ornithine and L-arginine and repression by ammonium

  • Leopoldo Naranjo
  • Mònica Lamas-Maceiras
  • Ricardo V. Ullán
  • Sonia Campoy
  • Fernando Teijeira
  • Javier Casqueiro
  • Juan F. MartínEmail author
Original Paper


The Penicillium chrysogenum oat1 gene, which encodes a class III ω-aminotransferase, was cloned and characterized. This enzyme converts lysine into 2-aminoadipic semialdehyde, and plays an important role in the biosynthesis of 2-aminoadipic acid, a precursor of penicillin and other β-lactam antibiotics. The enzyme is related to ornithine-5-aminotransferases and to the lysine-6-aminotransferases encoded by the lat genes found in bacterial cephamycin gene clusters. Expression of oat1 is induced by lysine, ornithine and arginine, and repressed by ammonium ions. AreA-binding GATA and GATT sequences involved in regulation by ammonium, and an 8-bp direct repeat associated with arginine induction in Emericella (Aspergillus nidulans and Saccharomyces cerevisiae, were found in the oat1 promoter region. Deletion of the oat1 gene resulted in the loss of ω-aminotransferase activity. The null mutants were unable to grow on ornithine or arginine as sole nitrogen sources and showed reduced growth on lysine. Complementation of the null mutant with the oat1 gene restored normal levels of ω-aminotransferase activity and the ability to grow on ornithine, arginine and lysine. The role of the oat1 gene in the biosynthesis of 2-aminoadipic acid is discussed.


Ω-Aminotransferase 2-Amino-adipic acid oat1 Penicillium chrysogenum Penicillin biosynthesis 



This work was supported by grants of the CICYT (BIO2000-1729-C02-02 and FIT-01000-2001-130). R.V. Ullán and F. Teijeira received fellowships from the Diputación de León and Junta de Castilla y León, respectively, L. Naranjo was granted a fellowship of the AECI Programme (Ministerio de Asuntos Exteriores, Madrid).


  1. Aharonowitz Y, Cohen G, Martín JF (1992) Penicillin and cephalosporin biosynthetic genes: structure, organization, regulation, and evolution. Annu Rev Microbiol 1992:461–495CrossRefGoogle Scholar
  2. Bhattacharjee JK (1985) α-Aminoadipate pathway for the biosynthesis of lysine in lower eukaryotes. Crit Rev Microbiol 12:131–151PubMedGoogle Scholar
  3. Cantoral JM, Díez B, Barredo JL, Álvarez E, Martín JF (1987) High frequency transformation of Penicillium chrysogenum. Biotechnology 5:494–497CrossRefGoogle Scholar
  4. Casqueiro J, Bañuelos O, Gutiérrez S, Hijarrubia MJ, Martín JF (1999) Intrachromosomal recombination in Penicillium chrysogenum: Gene conversion and deletion events. Mol Gen Genet 261:994–1000CrossRefPubMedGoogle Scholar
  5. Coque JJ, Liras P, Láiz L, Martín JL (1991) A gene encoding lysine 6-aminotransferase, which forms the β-lactam precursor α-aminoadipic acid, is located in the cluster of cephamycin biosynthetic genes in Nocardia lactamdurans. J Bacteriol 173:6258–6264PubMedGoogle Scholar
  6. Díez B, Álvarez E, Cantoral JM, Barredo JL, Martín JF (1987) Isolation and characterization of pyrG mutants of Penicillium chrysogenum by resistance to 5′-fluoroorotic acid. Curr Genet 12:277–282CrossRefGoogle Scholar
  7. Dzikowska A, Swianiewicz M, Talarczyk A, Wisniewska M, Goras M, Scazzocchio C, Weglenski P (1999) Cloning, characterisation and regulation of the ornithine transaminase (otaA) gene of Aspergillus nidulans. Curr Genet 35:118–126CrossRefPubMedGoogle Scholar
  8. Dzikowska A, Kacprzak M, Tomecki R, Koper M, Scazzocchio C, Weglenski P (2003) Specific induction and carbon/nitrogen repression of arginine catabolism gene of Aspergillus nidulans-functional in vivo analysis of the otaA promoter. Fungal Genet Biol 38:175–186CrossRefPubMedGoogle Scholar
  9. Empel J, Sitkiewicz I, Andrukiewicz A, Lasocki K, Borsuk P, Weglenski P (2001) arcA, the regulatory gene for the arginine catabolic pathway in Aspergillus nidulans. Mol Gen Genomics 266:591–597CrossRefGoogle Scholar
  10. Esmahan C, Álvarez E, Montenegro E, Martín JF (1994) Catabolism of lysine in Penicillium chrysogenum leads to formation of α-aminoadipic acid, a precursor of penicillin biosynthesis. Appl Enviroment Microbiol 60:1705–1710Google Scholar
  11. Fernandez FJ, Gutiérrez S, Velasco J, Montenegro E, Marcos AT, Martín JF (1994) Molecular characterization of three loss-of-function mutations in the isopenicillin N-acyltransferase gene (penDE) of Penicillium chrysogenum. J Bacteriol 176:4941–4948PubMedGoogle Scholar
  12. Fierro F, Gutiérrez S, Díez B, Martín JF (1993) Resolution of four large chromosomes in penicillin-producing filamentous fungi: the penicillin gene cluster is located on chromosome II (9.6 Mb) in Penicillium notatum and chromosome I (10.4 Mb) in Penicillium chrysogenum. Mol Gen Genet 241:573–578CrossRefPubMedGoogle Scholar
  13. Fierro F, Barredo JL, Díez B, Gutiérrez S, Fernández FJ, Martín JF (1995) The penicillin gene cluster is amplified in tandem repeats linked by conserved hexanucleotide sequences. Proc Natl Acad Sci USA 92:6200–6204PubMedCrossRefGoogle Scholar
  14. Fujii T, Narita T, Agematu H, Agata N, Isshiki K (2000) Characterization of L-lysine 6-aminotransferase and its structural gene from Flavobacterium lutescens IFO3084. J Biochem 128:391–397PubMedGoogle Scholar
  15. Guex N, Peitsch MC (1997) SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modelling. Electrophoresis 18:2714–2723CrossRefPubMedGoogle Scholar
  16. Gutiérrez S, Velasco J, Marcos AT, Fernández FJ, Fierro F, Barredo JL, Díez B, Martín JF (1997) Expression of the cefG gene is limiting for cephalosporin biosynthesis in Acremonium chrysogenum. Appl Microbiol Biotechnol 48:606–614CrossRefPubMedGoogle Scholar
  17. Heimberg H, Boyen A, Crabeel M, Glansdorff N (1990) Escherichia coli and Saccharomyces cerevisiae acetylornithine aminotransferase: evolutionary relationship with ornithine aminotransferases. Gene 90:69–78CrossRefPubMedGoogle Scholar
  18. Ijlst L, de Kromme I, Oostheim W, Wanders RJ (2000) Molecular cloning and expression of human L-pipecolate oxidase. Biochem Biophys Res Commun 270:1101–1105CrossRefPubMedGoogle Scholar
  19. Kern BA, Hendlin D, Inamine E (1980) L-lysine ε-aminotransferase involved in cephamycin C synthesis in Streptomyces lactamdurans. Antimicrob Agents Chemother 17:679–685PubMedGoogle Scholar
  20. Kumar S, Tamura K, Jakobsen IB, Nei M (2001) MEGA2: Molecular Evolutionary Genetics Analysis software. Bioinformatics 17:1244–1245 ( Scholar
  21. Liu G, Casqueiro J, Bañuelos O, Cardoza RE, Gutiérrez S, Martín JF (2001) Targeted inactivation of the mecB gene encoding cystathionine-Y-lyase shows that the transsulfuration pathway is required for high level cephalosporin biosynthesis in Acremonium chrysogenum C10 but not for methionine induction of the cephalosporin genes. J Bacteriol 183:1765–1772CrossRefPubMedGoogle Scholar
  22. Madduri K, Stuttard C, Vining JC (1991) Cloning and location of a gene governing lysine epsilon-aminotransferase, an enzyme initiating β-lactam biosynthesis in Streptomyces sp. J Bacteriol 173:985–988PubMedGoogle Scholar
  23. Martín JF (1998) New aspects of genes and enzymes for β-lactam antibiotic biosynthesis. Appl Microbiol Biotechnol 50:1–15CrossRefPubMedGoogle Scholar
  24. Martín de Valmaseda E, Campoy S, Naranjo L, Casqueiro J, Martín JF (2005) Lysine is catabolized to 2-aminoadipic acid in Penicillium chrysogenum by an ω-aminotransferase and to saccharopine by a lysine α-ketoglutarate reductase: characterization of the ω-aminotransferase. Mol Genet Genomics (accompanying paper)Google Scholar
  25. Messenguy F, Vierendeels F, Scherens B, Dubois E (2000) In Saccharomyces cerevisiae, Expression of arginine catabolic genes CAR1 and CAR2 in response to exogenous nitrogen availability is mediated by the Ume6 (CargRI)-Sin3 (CargRII)-Rpd3 (CargRIII) complex. J Bacteriol 182:3158–3164CrossRefPubMedGoogle Scholar
  26. Naranjo L, Martín de Valmaseda E, Bañuelos O, López P, Riaño J, Casqueiro J, Martín JF (2001) Conversion of pipecolic acid into lysine in Penicillium chrysogenum requires pipecolate oxidase and saccharopine reductase: characterization of the lys7 gene encoding saccharopine reductase. J Bacteriol 183:7165–7172CrossRefPubMedGoogle Scholar
  27. Naranjo L, Martín de Valmaseda E, Casqueiro J, Ullán RV, Lamas M, Bañuelos O, Martín JF (2004) Inactivation of the lys7 gene, encoding saccharopine reductase in Penicillium chrysogenum, leads to accumulation of the secondary metabolite precursors piperideine-6-carboxylic acid and pipecolic acid from α-aminoadipic acid. Applied Environ Microbiol 70:1031–1039CrossRefGoogle Scholar
  28. Pérez-Llarena FJ, Rodríguez-García A, Enguita FJ, Martín JF, Liras P (1998) The pcd gene encoding piperideine-6-carboxylate dehydrogenase involved in biosynthesis of α-aminoadipic acid is located in the cephamycin cluster of Streptomyces clavuligerus. J Bacteriol 180:4753–4756PubMedGoogle Scholar
  29. Romero J, Martín JF, Liras P, Demain AL, Rius N (1997) Partial purification, characterization and nitrogen regulation of the lysine ε-aminotransferase of Streptomyces clavuligerus. J Ind Microbiol Biotechnol 18:241–246CrossRefPubMedGoogle Scholar
  30. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring HarborGoogle Scholar
  31. Schwede T, Kopp J, Guex N, Peitsch MC (2003) SWISS-MODEL: an automated protein homology-modeling server. Nucleic Acids Res 31:3381–3385 ( Scholar
  32. Shen BW, Hennig M, Hohenester E, Jansonius JN, Schirmer T (1998) Crystal structure of human recombinant ornithine aminotransferase. J Mol Biol 277:81–102CrossRefPubMedGoogle Scholar
  33. Sim KL, Perry D (1997) Analysis of swainsonine and its early metabolic presursors in cultures of Metarhizium anisopliae. Glycoconj J 14:661–668CrossRefPubMedGoogle Scholar
  34. Watanabe N, Yonaha K, Sakabe K, Sakabe N, Aibara S, Morita Y (1991) Crystal structure of ω-amino acid:pyruvate aminotransferase. In: Fukui T, Kagamiyama H, Soda K, Wada H (eds) Enzymes dependent on pyridoxal phosphate and other carbonyl compounds as cofactors. Pergamon Press, Oxford, pp 121–124Google Scholar
  35. Wickwire BM, Harris CM, Harris TM, Broquist HP (1990) Pipecolic acid biosynthesis in Rhizoctonia leguminicola. I. The lysine saccharopine, delta 1-piperideine-6-carboxylic acid pathway. J Biol Chem 265:14742–14747PubMedGoogle Scholar
  36. Yasuda M, Tanizawa K, Misono H, Toyama S, Soda K (1981) Properties of crystalline L-ornithine: α-ketoglutarate delta-aminotransferase from Bacillus sphaericus. J Bacteriol 148:43–50PubMedGoogle Scholar
  37. Yonaha K, Nishie M, Aibara S (1992) The primary structure of omega-amino acid:pyruvate aminotransferase. J Biol Chem 267:12506–12510PubMedGoogle Scholar
  38. Yu H, Serpe HYE, Romero J, Coque JJR, Maeda K, Oelgeschläger M, Hintermann G, Liras P, Martín JF, Demain AL, Piret J (1994) Possible involvement of the lysine E-aminotransferase gene (lat) in the expression of the genes encoding ACV synthetase (pcbAB) and isopenicillin N synthase (pcbC) in Streptomyces clavuligerus. Microbiology 140:3367–3377PubMedCrossRefGoogle Scholar
  39. Zhu X, Tang G, Galili G (2000) Characterization of the two saccharopine dehydrogenase isozymes of lysine catabolism encoded by the single composite AtLKR/SDH locus of Arabidopsis. Plant Physiol 124:1363–1371CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Leopoldo Naranjo
    • 1
    • 3
  • Mònica Lamas-Maceiras
    • 2
  • Ricardo V. Ullán
    • 1
  • Sonia Campoy
    • 1
  • Fernando Teijeira
    • 1
  • Javier Casqueiro
    • 1
    • 2
  • Juan F. Martín
    • 1
    • 2
    Email author
  1. 1.Instituto de Biotecnología de León (INBIOTEC)Parque Científico de LeónLeónSpain
  2. 2.Área de Microbiología, Fac. CC. Biológicas y AmbientalesUniversidad de LeónLeónSpain
  3. 3.Centro de BiotecnologíaInstituto de Estudios Avanzados (IDEA)CaracasVenezuela

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