Applied Microbiology and Biotechnology

, Volume 77, Issue 6, pp 1305–1316

Expression of Clostridium acetobutylicum butanol synthetic genes in Escherichia coli

Authors

  • Masayuki Inui
    • Research Institute of Innovative Technology for the Earth
  • Masako Suda
    • Research Institute of Innovative Technology for the Earth
  • Sakurako Kimura
    • Research Institute of Innovative Technology for the Earth
  • Kaori Yasuda
    • Research Institute of Innovative Technology for the Earth
  • Hiroaki Suzuki
    • Research Institute of Innovative Technology for the Earth
  • Hiroshi Toda
    • Research Institute of Innovative Technology for the Earth
  • Shogo Yamamoto
    • Research Institute of Innovative Technology for the Earth
  • Shohei Okino
    • Research Institute of Innovative Technology for the Earth
  • Nobuaki Suzuki
    • Research Institute of Innovative Technology for the Earth
    • Research Institute of Innovative Technology for the Earth
Applied Genetics and Molecular Biotechnology

DOI: 10.1007/s00253-007-1257-5

Cite this article as:
Inui, M., Suda, M., Kimura, S. et al. Appl Microbiol Biotechnol (2008) 77: 1305. doi:10.1007/s00253-007-1257-5

Abstract

A recombinant butanol pathway composed of Clostridium acetobutylicum ATCC 824 genes, thiL, hbd, crt, bcd-etfB-etfA, and adhe1 (or adhe) coding for acetyl-CoA acetyltransferase (THL), β-hydroxybutyryl-CoA dehydrogenase (HBD), 3-hydroxybutyryl-CoA dehydratase (CRT), butyryl-CoA dehydrogenase (BCD), butyraldehyde dehydrogenase (BYDH), and butanol dehydrogenase (BDH), under the tac promoter control was constructed and was introduced into Escherichia coli. The functional expression of these six enzymes was proved by demonstrating the corresponding enzyme activities using spectrophotometric, high performance liquid chromatography and gas chromatography analyses. The BCD activity, which was not detected in E. coli previously, was shown in the present study by performing the procedure from cell extract preparation to activity measurement under anaerobic condition. Moreover, the etfA and etfB co-expression was found to be essential for the BCD activity. In the case of BYDH activity, the adhe gene product was shown to have higher specificity towards butyryl-CoA compared to the adhe1 product. Butanol production from glucose was achieved by the highly concentrated cells of the butanologenic E. coli strains, BUT1 with adhe1 and BUT2 with adhe, under anaerobic condition, and the BUT1 and BUT2 strains were shown to produce 4 and 16-mM butanol with 6- and 1-mM butyrate as a byproduct, respectively. This study reports the novel butanol production by an aerobically pregrown microorganism possessing the genes of a strict anaerobe, Clostridium acetobutylicum.

Keywords

Butanol productionClostridium acetobutylicumEscherichia coliBiofuel

Copyright information

© Springer-Verlag 2007