Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Research Paper
  • Published:

Construction of a 7-Aminocephalosporanic Acid (7ACA) Biosynthetic Operon and Direct Production of 7ACA in Acremonium Chrysogenum

Abstract

We have used cDNA encoding D-amino acid oxidase, and genomic DNA encoding cephalosporin acylase from Fusarium solani and Pseudomonas diminuta, respectively, to construct a novel hybrid 7-aminocephalosporanic acid (7ACA) biosynthetic operon under the control of regulatory elements from the alkaline protease gene of Acremonium chrysogenum. Transformants of A. chrysogenum BC2116, a high cephalosporin-producing strain, containing this operon, synthesized and secreted low levels of 7ACA. Although the amounts are not yet commercially significant, this represents the first microbial production of 7ACA and demonstrates the feasibility of introducing new biosynthetic capabilities into industrial microorganisms by combining fungal and bacterial genes.

This is a preview of subscription content, access via your institution

Access options

Buy this article

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Bunnel, C.A., Luke, W.D. and Perry, F.M. 1986. Industrial manufacture of cephalosporins, p.255–284. In: Beta-lactam Antibiotics for Clinical Use. S.F. Queener, J.A. Webber, and S.W. Queener (Eds.). Marcel Dekker, New York.

  2. Tsuzuki, K., Komatsu, K.-I., Ichikawa, S. and Shibuya, Y. 1989. Enzymatic synthesis of 7-aminocephalosporanic acid (7-ACA). Nippon Nogeikagaku Kaishi 63: 1847–1853.

    Article  CAS  Google Scholar 

  3. Malpartida, F. and Hopwood, D.A. 1984. Molecular cloning of the whole biosynthetic pathway of a Streptomyces antibiotic and its expression in a heterologous host. Nature 309: 462–464.

    Article  CAS  Google Scholar 

  4. Chen, C.W., Lin, H.-F., Kuo, C.L., Tsai, H.-L. and Tsai, J.F.-Y. 1988. Cloning and expression of a DNA sequence conferring cephamycin C production. Bio/Technology 6: 1222–1224.

    Article  CAS  Google Scholar 

  5. Smith, D.J., Burnham, M.K.R., Edward, J., Earl, A.J. and Turner, G. 1990. Cloning and heterologous expression of the penicillin biosynthetic gene cluster from Penicillium chrysogenum. Bio/Technology 8: 39–41.

    CAS  PubMed  Google Scholar 

  6. Isogai, T., Ono, H. and Kojo, H. 1990. D-Amino acid oxidase. Eur. Pat. Appl. EP 364,275 (18 Apr 1990).

  7. Isogai, T., Ono, H., Ishitani, Y., Kojo, H., Ueda, Y. and Kohsaka, M. 1990. Structure and expression of cDNA for D-amino acid oxidase active against cephalosporin C from Fusarium solani. J. Biochem. 108: 1063–1069.

    Article  CAS  Google Scholar 

  8. Szwajcer, E. and Mosbach, K. 1985. Isolation and partial characterization of a D-amino acid oxidase active against cephalosporin C from the yeast Trigonopsis variabilis. Biotechnology Letters 7: 1–7.

    Article  CAS  Google Scholar 

  9. Matsuda, A., Matsuyama, K., Yamamoto, K., Ichikawa, S. and Komatsu, K.-I. 1987. Cloning and characterization of the genes for two distinct cephalosporin acylases from a Pseudomonas strain. J. Bacteriol. 169: 5815–5820.

    Article  CAS  Google Scholar 

  10. Matsuda, A., Toma, K. and Komatsu, K.-I. 1987. Nucleotide sequences of the genes for two distinct cephalosporin acylases from a Pseudomonas strain. J. Bacteriol. 169: 5821–5826.

    Article  CAS  Google Scholar 

  11. Isogai, T., Fukagawa, M., Kojo, H., Kohsaka, M., Aoki, H. and Imanaka, H. 1991. Cloning and nucleotide sequence of the complementary and genomic DNAs for the alkaline protease from Acremonium chrysogenum. Agric. Biol. Chem. In press.

    Google Scholar 

  12. Yagi, J., Yano, T., Kubochi, Y., Hattori, S., Ohashi, M., Sakai, H., Jomon, K. and Ajisaka, M. 1972. Studies on alkaline proteinase from Cephalosporium. J. Ferment. Technol. 50: 592–599.

    CAS  Google Scholar 

  13. Skatrud, P.L., Queener, S.W., Carr, L.G. and Fisher, D.L. 1987. Efficient integrative transformation of Cephalosporium acremonium. Curr. Genet. 12: 337–348.

    Article  CAS  Google Scholar 

  14. Skatrud, P.L., Tietz, A.J., Ingolia, T.D., Cantwell, C.A., Fisher, D.L., Chapman, J.L. and Queener, S.W. 1989. Use of recombinant DNA to improve production of cephalosporin C by Cephalosporium acremonium. Bio/Technology 7: 477–485.

    CAS  Google Scholar 

  15. Gritz, L. and Davies, J. 1983. Plasmid-encoded hygromycin B resistance: the sequence of hygromycin B phosphotransferase gene and its expression in Escherichia coli and Saccharomyces cerevisiae. Gene 25: 179–188.

    Article  CAS  Google Scholar 

  16. van den Elzen, P.J.M., Townsend, J., Lee, K.Y. and Bedbrook, J.R. 1985. A chimaeric hygromycin resistance gene as a selectable marker in plant cells. Plant Molecular Biology 5: 299–302.

    Article  CAS  Google Scholar 

  17. Takeshita, S., Sato, M., Toba, M., Masahashi, W. and HashimotoGotoh, T. 1987. High-copy-number and low-copy-number plasmid vectors for lacZα-complementation and chloramphenicol- or kanamycin-resistance selection. Gene 61: 63–74.

    Article  CAS  Google Scholar 

  18. Shen, Y.-Q., Wolfe, S. and Demain, A.L. 1986. Levels of isopenicillin N synthetase and deacetoxycephalosporin C synthetase in Cephalosporium acremonium producing high and low levels of cephalosporin C. Bio/Technology 4: 61–64.

    CAS  Google Scholar 

  19. Maniatis, T., Fritsch, E.F. and Sambrook, J. 1982. Molecular Cloning. A Laboratory Manual. Cold Spring Harbor, New York.

    Google Scholar 

  20. Isogai, T., Yoshida, M., Tanaka, M. and Aoki, H. 1987. Transformation of Acremonium chrysogenum and Saccharomyces cerevisiae using an antibiotic resistance marker. Agric. Biol. Chem. 51: 2321–2329.

    CAS  Google Scholar 

  21. Samson, S.M., Belagaje, R., Blankenship, D.T., Chapman, J.L., Perry, D., Skatrud, P.L., VanFrank, R.M., Abraham, E.P., Baldwin, J.E., Queener, S.W. and Ingolia, T.D. 1985. Isolation, sequence determination and expression in Escherichia coli of the isopenicillin N synthetase gene from Cephalosporium acremonium. Nature 318: 191–194.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Isogai, T., Fukagawa, M., Aramori, I. et al. Construction of a 7-Aminocephalosporanic Acid (7ACA) Biosynthetic Operon and Direct Production of 7ACA in Acremonium Chrysogenum. Nat Biotechnol 9, 188–191 (1991). https://doi.org/10.1038/nbt0291-188

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt0291-188

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing