Applied Microbiology and Biotechnology

, Volume 56, Issue 1, pp 144–149

Construction of an expression vector for propionibacteria and its use in production of 5-aminolevulinic acid by Propionibacterium freudenreichii

  •  P. Kiatpapan
  •  Y. Murooka
Original Paper

DOI: 10.1007/s002530100603

Cite this article as:
Kiatpapan, P. & Murooka, Y. Appl Microbiol Biotechnol (2001) 56: 144. doi:10.1007/s002530100603


Several promoters from Propionibacterium freudenreichii subsp. shermanii were isolated using a promoter probe vector, pCVE1, containing the Streptomyces cholesterol oxidase gene (choA) as a reporter gene. Three of four promoters isolated exhibiting a strong activity in Escherichia coli also expressed a strong activity in P. freudenreichii subsp. shermanii IFO12426. Using two promoters with a strong activity and a previously constructed shuttle vector, pPK705, shuttling between E. coli and Propionibacterium, we constructed expression vectors for propionibacteria. To overproduce 5-aminolevulinic acid (ALA), which is the first intermediate in the synthesis of porphyrins, the ALA synthase gene (hemA) from Rhodobacter sphaeroides was recombined with the expression vectors. The activity of ALA synthase in the recombinant P. freudenreichii subsp. shermanii increased about 70-fold that in the strain without a vector. The recombinant Propionibacterium produced ALA at a maximum concentration of 8.6 mM in the absence of levulinic acid, an inhibitor of ALA dehydratase, with 1% glucose as a carbon source. The recombinant P. freudenreichii accumulated 18.8 mmol/g cells ALA in the presence of 1 mM levulinic acid and 30 mM glycine. The construction of an efficient expression vector will facilitate genetic studies of a vitamin B12 producer, Propionibacterium.

Copyright information

© Springer-Verlag 2001

Authors and Affiliations

  •  P. Kiatpapan
    • 2
  •  Y. Murooka
    • 1
  1. 1.Department of Biotechnology, Graduate School of Engineering, Osaka University, Yamada-oka, Suita, Osaka 565-0871, Japan
  2. 2.Department of Biochemistry, Faculty of Science, Rangsit University, Patumtani 12000, Thailand