Heterologous expression of secreted biologically active human interleukin-10 in Bifidobacterium breve

Construction of Bifidobacterium breve capable of production of secreted biologically active human interleukin-10 (hIL-10) is described. ORF coding for full-length mature human interleukin-10 was cloned into a series of expression vectors. This resulted in generation of translational fusions between...

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Bibliographic Details
Published in:Archives of microbiology 2010-09, Vol.192 (9), p.769-774
Main Authors: Khokhlova, E. V, Efimov, B. A, Kafarskaia, L. I, Shkoporov, A. N
Format: Article
Language:English
Subjects:
Bifidobacterium
Bifidobacterium - genetics
Bifidobacterium - metabolism
Biochemistry
Biomedical and Life Sciences
Biotechnology
Bowel disease
Cell Biology
Cloning
Cloning, Molecular
Culture Media
Cytokines
E coli
Ecology
Gene Expression Regulation, Bacterial
Genes
Genetic Vectors
Growth conditions
HT29 Cells
Humans
Industrial Microbiology
inflammatory bowel disease
interleukin-10
Interleukin-10 - biosynthesis
Life Sciences
Microbial Ecology
Microbiology
Microbiota
Original Paper
Peptides
Plasmids
Promoter Regions, Genetic
Recombinant Fusion Proteins - biosynthesis
Vector
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Summary:Construction of Bifidobacterium breve capable of production of secreted biologically active human interleukin-10 (hIL-10) is described. ORF coding for full-length mature human interleukin-10 was cloned into a series of expression vectors. This resulted in generation of translational fusions between hIL-10 and signal peptides sequences derived from Bifidobacterium breve genes sec2, apuB and B. adolescentis gene amyB under the control of constitutively active bifidobacterial promoter. We have shown that fusion to amyB signal peptide resulted in highest expression level of hIL-10 at the mRNA and protein level. Secreted hIL-10 was highly unstable in bifidobacterial culture supernatants in standard growth conditions. However, incubation of stationary cultures in buffered tissue culture medium resulted in production of stable biologically active hIL-10, albeit in low amounts (1.9 ng/ml).
ISSN:0302-8933
1432-072X
DOI:10.1007/s00203-010-0606-4