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

Biotechnology Letters volume 30, Article number: 1403 (2008) Cite this article

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.

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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.

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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
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  2. Toms C. Joseph
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  3. Nirmala Thampuran
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  4. Roswin James
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  5. Kesavan Ashok Kumar
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  6. Chinnusamy Viswanathan
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  7. Kailash C. Bansal
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Correspondence to Toms C. Joseph.

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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 (2008). https://doi.org/10.1007/s10529-008-9688-3

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

Keywords

  • Bacillus halodurans
  • Compatible solutes
  • Ectoine genes
  • Osmoregulation
  • Salt stress