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J Biosci Bioeng. 2015 May;119(5):515-20. doi: 10.1016/j.jbiosc.2014.09.024. Epub 2014 Oct 29.

Cloning and heterologous expression of the ftfCNC-2(1) gene from Weissella confusa MBFCNC-2(1) as an extracellular active fructansucrase in Bacillus subtilis.

Author information

1
Pharmaceutical Biotechnology Research Group, Faculty of Pharmacy, Universitas Indonesia, Kampus UI Depok, Depok 16424, Indonesia. Electronic address: amarila.malik@ui.ac.id.
2
Pharmaceutical Biotechnology Research Group, Faculty of Pharmacy, Universitas Indonesia, Kampus UI Depok, Depok 16424, Indonesia.
3
Graduate School of Biological Sciences, Nara Institute of Science and Technology (NAIST), 8916-5 Takayama, Ikoma, Nara 630-0101, Japan.

Abstract

Fructan-exopolysaccharides (fructan-EPS) (inulin and levan) and their oligosaccharides (fructooligosaccharides, FOS) have drawn considerable interest in the food and pharmaceutical industries. EPS-producing lactic acid bacteria have been reported to produce β-fructans (inulin and levan), as well as α-glucans, by the function of sucrase enzymes, i.e., fructansucrase and glucansucrase. A fructansucrase ftfCNC-2(1) gene from Weissella confusa strain MBFCNC-2(1) was previously cloned in Escherichia coli. In this study, we aimed to express the ftf[CNC-2(1)] gene in Bacillus subtilis to obtain the active form of the extracellular recombinant protein FTF[CNC-2(1)]. This cloning was achieved by inserting the gene in-fusion with the signal sequence of the B. subtilis subtilisin E. SDS-polyacrylamide gel electrophoresis analysis and in situ activity assay with Periodic Acid-Schiff staining revealed that the recombinant FTF[CNC-2(1)] was successfully expressed as an extracellular protein from B. subtilis DB403 in its active form, which was confirmed using sucrose and raffinose.

KEYWORDS:

Bacillus subtilis; Fructansucrase; Fructosyltransferase; Inulosucrase; Weissella confusa; ftf

PMID:
25454699
DOI:
10.1016/j.jbiosc.2014.09.024
[Indexed for MEDLINE]

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