Skip to main content
SpringerLink
Log in

High Soluble Expression of d-Amino Acid Oxidase in Escherichia coli Regulated by a Native Promoter

  • Published:
Applied Biochemistry and Biotechnology Aims and scope Submit manuscript

Abstract

To express high-active soluble d-amino acid oxidase (DAAO), a constitutive plasmid that is regulated by a native hydantoinase promoter (PHase), was constructed. A d-amino acid oxidase gene (dao) was ligated with the PHase and cloned into pGEMKT to constitutively express protein of DAAO without the use of any inducer such as isopropyl β-d-1-thiogalactopyranoside which is poisonous to the cells and environment. The ribosome binding site region, host strain, and fermentation conditions were optimized to increase the expression level. When cultivated in a 5-m3 fermenter, the enzyme activity of JM105/pGEMKT-R-DAAO grown at 37 °C was found to be 32 U/mL and increase 16-fold over cells of BL21(DE3)/pET-DAAO grown at 28 °C. These results indicate the success of our approaches to overproducing DAAO in soluble form in Escherichia coli.

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

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  1. Upadhya, R., Nagajyothi, H., & Bhat, S. D. (1999). Process Biochemistry, 35, 7–13. doi:10.1016/S0032-9592(99)00025-4.

    Article  CAS  Google Scholar 

  2. Patel, R. N. (2001). Biomolecular Engineering, 17, 167–182. doi:10.1016/S1389-0344(01)00068-5.

    Article  CAS  Google Scholar 

  3. Pilone, M. S., & Pollegioni, L. (2002). Biocatalysis and Biotransformation, 20, 145–159. doi:10.1080/10242420290020679.

    Article  CAS  Google Scholar 

  4. Pollegioni, L., Caldinelli, L., Molla, G., Sacchi, S., & Pilone, M. S. (2004). Biotechnology Progress, 20, 467–473. doi:10.1021/bp034206q.

    Article  CAS  Google Scholar 

  5. Tishkov, V. I., & Khoronenkova, S. V. (2005). Biochemistry, 70, 40–54.

    CAS  Google Scholar 

  6. Setoyama, C., Miura, R., Nishina, Y., Shiga, K., Mizutani, H., Miyahara, I., et al. (1996). Journal of Biochemistry, 119, 1114–1117.

    CAS  Google Scholar 

  7. Isogai, T., Ono, H., Ishitani, Y., Kojo, H., Ueda, Y., & Kohsaka, M. (1990). Journal of Biochemistry, 108, 1063–1069.

    CAS  Google Scholar 

  8. Molla, G., Vegezzi, C., Pilone, M. S., & Pollegioni, L. (1998). Protein Expression and Purification, 14, 289–294. doi:10.1006/prep.1998.0956.

    Article  CAS  Google Scholar 

  9. Harris, C. M., Molla, G., Pilone, M. S., & Pollegioni, L. (1999). The Journal of Biological Chemistry, 274, 36233–36240. doi:10.1074/jbc.274.51.36233.

    Article  CAS  Google Scholar 

  10. Alonso, J., Barredo, J. L., Armisen, P., Diez, B., Salto, F., Guisan, J. M., et al. (1999). Enzyme and Microbial Technology, 25, 88–95. doi:10.1016/S0141-0229(99)00019-8.

    Article  CAS  Google Scholar 

  11. Isogai, A., Kimura, H., Reichert, A., Schorgendorfer, K., Nikaido, K., Tohda, H., et al. (2002). Biotechnology and Bioengineering, 80, 22–32. doi:10.1002/bit.10345.

    Article  Google Scholar 

  12. Luo, H., Tong, Y. Z., Li, Q., Yu, H. M., & Shen, Z. Y. (2004). Acta Microbiologica Sinica, 44, 336–339.

    CAS  Google Scholar 

  13. Lee, Y. H., Chu, W. S., & Hsu, W. H. (1994). Biotechnology Letters, 16, 467–472. doi:10.1007/BF01023327.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Chemical Engineering, Institute of Biochemical Engineering, Tsinghua University, Beijing, 100084, People’s Republic of China

    Yangqiu Liu, Qiang Li, Hongyu Zhu & Jichu Yang

Authors
  1. Yangqiu Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Qiang Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hongyu Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jichu Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Qiang Li.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Liu, Y., Li, Q., Zhu, H. et al. High Soluble Expression of d-Amino Acid Oxidase in Escherichia coli Regulated by a Native Promoter. Appl Biochem Biotechnol 158, 313–322 (2009). https://doi.org/10.1007/s12010-008-8325-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12010-008-8325-x

Keywords

Search

Navigation