Skip to main content
SpringerLink
Log in

Cloning and heterologous expression of ectoine biosynthesis genes from Bacillus halodurans in Escherichia coli

  • Original Research Paper
  • Published:
Biotechnology Letters Aims and scope Submit manuscript

Abstract

The genes involved in the biosynthetic pathway of ectoine (2-methyl-1,4,5,6-tetrahydropyrimidine-4-carboxylic acid) from Bacillus halodurans were cloned as an operon and expressed in E. coli. Analysis of the deduced ectoine biosynthesis cluster amino acid sequence revealed that the ectoine operon contain 2,389 bp, encoded by three genes; ectA, ectB and ectC that encode proteins of 189, 427 and 129 amino acids with deduced molecular masses of 21,048, 47,120 and 14,797 Da respectively. Extracts of induced cells showed two bands at 41 kDa and 17 kDa, possibly corresponding to the products of the later two genes. However the expression of ectA gene could not be ascertained by SDS-PAGE. The activity of the ectA protein was confirmed by an acylation assay. The transgenic E. coli accumulated upto 4.6 mg ectoine/l culture. This is the first report of an engineered E. coli strain carrying the ectoine genes of the alkaliphilic bacterium, B. halodurans.

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

Similar content being viewed by others

References

  • Ausubel FM, Brent R, Kingston RE, Moore DD, Scidman JG, Smith JA, Strichi K (1994) Current protocols in molecular biology. John Wiley and sons, New York

    Google Scholar 

  • Canovas D, Vargas C, Calderon MI, Ventosa A, Nieto JJ (1998) Characterization of the genes for the biosynthesis of the compatible solute ectoine in the moderately halophilic bacterium Halomonas elongata DSM 3043. Syst Appl Microbiol 21:487–497

    PubMed  CAS  Google Scholar 

  • Csonka LN, Hanson AD (1991) Prokaryotic osmoregulation: genetics and physiology. Annu Rev Microbiol 45:569–606

    Article  PubMed  CAS  Google Scholar 

  • Galinski EA (1995) Osmoadaptation in bacteria. Adv Microb Physiol 37:273–328

    CAS  Google Scholar 

  • Galinski EA, Pfeiffer HP, Truper HG (1985) 1,4,5,6-Tetrahydro-2-methyl-4-pyrimidinecarboxylic acid. A novel cyclic amino acid from halophilic phototrophic bacteria of the genus Ectothiorhodospira. Eur J Biochem 149:135–139

    Article  PubMed  CAS  Google Scholar 

  • Horikoshi K (1999) Alkaliphiles: some applications of their products for biotechnology. Microbiol Mol Biol Rev 63:735–750

    PubMed  CAS  Google Scholar 

  • Kempf B, Bremer E (1998) Uptake and synthesis of compatible solutes as microbial stress responses to high-osmolality environments. Arch Microbiol 170:319–330

    Article  PubMed  CAS  Google Scholar 

  • Kuhlmann AU, Bremer E (2002) Osmotically regulated synthesis of the compatible solute ectoine in Bacillus pateurii and related Bacillus spp. Appl Environ Microbiol 68:772–783

    Article  PubMed  CAS  Google Scholar 

  • Louis P, Galinski EA (1997) Characterization of genes for the biosynthesis of the compatible solute ectoine from Marinococcus halophilus and osmoregulated expression in Escherichia coli. Microbiology 143:1141–1149

    Article  PubMed  CAS  Google Scholar 

  • Nagata S, Wang YB (2001) Accumulation of ectoine in the halotolerant Brevibacterium spp. JCM 6894. J Biosci Bioeng 91:288–293

    Article  PubMed  CAS  Google Scholar 

  • Ono H, Sawada K, Khunajakr N, Tao T, Yamamoto M, Hiramoto M, Shinmyo A, Takano M, Murooka Y (1999) Characterization of biosynthetic enzymes for ectoine as a compatible solute in a moderately halophilic eubacterium, Halomonas elongata. J Bacteriol 181:91–99

    PubMed  CAS  Google Scholar 

  • Schubert T, Maskow T, Benndorf D, Harms H, Breuer U (2007) Continuous synthesis and excreation of the compatible solute ectoine by a transgenic, nonhalophilic bacterium. Appl Environ Microbiol 73:3343–3347

    Article  PubMed  CAS  Google Scholar 

  • Sleator RD, Hill C (2002) Bacterial osmoadaptation: the role of osmolytes in bacterial stress and virulence. FEMS Microbiol Rev 26:49–71

    Article  PubMed  CAS  Google Scholar 

  • Takami H, Horikoshi K (1999) Reidentification of facultatively alkaliphilic Bacillus spp. C-125 to Bacillus halodurans. Biosci Biotech Biochem 63:943–945

    Article  CAS  Google Scholar 

  • Vargas C, Jebbar M, Carrasco R, Blanco C, Calderon MI, IgIesias-Guerra F, Nieto JJ (2006) Ectoines as compatible solutes and carbon and energy sources for the halophilic bacterium Chromohalobacter salexigens. J Appl Microbiol 100:98–107

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

Authors are grateful to the Director, Central Institute of Fisheries Technology (CIFT), Cochin for providing the necessary facilities to carry out this research work and Indian Council for Agricultural Research, New Delhi, India for financial assistance.

Author information

Authors and Affiliations

  1. Microbiology, Fermentation and Biotechnology Division, Central Institute of Fisheries Technology, Cochin, 682 029, Kerala, India

    Lawrance Anbu Rajan, Toms C. Joseph, Nirmala Thampuran & Roswin James

  2. Quality Assurance and Management Division, Central Institute of Fisheries Technology, Cochin, 682 029, Kerala, India

    Kesavan Ashok Kumar

  3. Water Technology Centre, Indian Agricultural Research Institute, New Delhi, 110 012, India

    Chinnusamy Viswanathan

  4. National Research Centre on Plant Biotechnology, Indian Agricultural Research Institute, New Delhi, 110 012, India

    Kailash C. Bansal

Authors
  1. Lawrance Anbu Rajan
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Toms C. Joseph
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Nirmala Thampuran
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Roswin James
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Kesavan Ashok Kumar
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Chinnusamy Viswanathan
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Kailash C. Bansal
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Toms C. Joseph.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Anbu Rajan, L., Joseph, T.C., Thampuran, N. et al. Cloning and heterologous expression of ectoine biosynthesis genes from Bacillus halodurans in Escherichia coli . Biotechnol Lett 30, 1403–1407 (2008). https://doi.org/10.1007/s10529-008-9688-3

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10529-008-9688-3

Keywords

Search

Navigation