Production of human basic fibroblast growth factor (FGF-2) in Bifidobacterium breve using a series of novel expression/secretion vectors

Four E. coli-Bifidobacterium shuttle vectors were constructed using Bifidobacterium plasmids, pB44 and pB80. The vectors carry two bifidobacterial promoters, a signal peptide-encoding sequence, sec2, of Bifidobacterium breve, and a transcriptional terminator from hup gene of Bifidobacterium longum....

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Bibliographic Details
Published in:Biotechnology letters 2008-11, Vol.30 (11), p.1983-1988
Main Authors: Shkoporov, A. N, Efimov, B. A, Khokhlova, E. V, Kafarskaia, L. I, Smeianov, V. V
Format: Article
Language:English
Subjects:
Applied Microbiology
Bifidobacterium - genetics
Bifidobacterium breve
Bifidobacterium longum
Biochemistry
Biomedical and Life Sciences
Biotechnology
Blotting, Western
E coli
Fibroblast Growth Factor 2 - genetics
Fibroblast Growth Factor 2 - metabolism
Genetic Vectors - genetics
Genomics
Gram-positive bacteria
Humans
Life Sciences
Microbiology
Original Research Paper
Pharmacology
Plasmids - genetics
Recombinant Proteins - metabolism
Toxicology
Transformation, Genetic
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Summary:Four E. coli-Bifidobacterium shuttle vectors were constructed using Bifidobacterium plasmids, pB44 and pB80. The vectors carry two bifidobacterial promoters, a signal peptide-encoding sequence, sec2, of Bifidobacterium breve, and a transcriptional terminator from hup gene of Bifidobacterium longum. Functionality of the constructs were tested using human FGF-2 gene. The expression of FGF-2 was detected by Western blotting in B. breve transformed with three of the vectors. The highest amount of FGF-2 was produced upon transformation with pESH86, which is a pB80-based plasmid carrying FGF-2 under control of a hup promoter (Phup). Similarly, the level of FGF-2 mRNA transcribed from pESH86 was approximately threefold higher, 882 ± 70 AU (arbitrary units), when compared to those transcribed from pB44-based pESH46 (Phup) (289 ± 65 AU) and pESH47 (Pgap) (282 ± 37 AU). These results suggest the vectors have the potential for production of exported fusion proteins in bifidobacteria and the expression levels can be regulated through the employment of different bifidobacterial promoters and/or replicons.
ISSN:0141-5492
1573-6776
DOI:10.1007/s10529-008-9772-8