Molecular cloning and in silico characterization of fructansucrase gene from weissella confusa MBFCNC-2(1) isolated from local beverage

Fructan-type exopolysaccharides (EPS) (inulin and levan) and their oligosaccharides (fructooligosaccharides/FOS) are of great interest to the food and pharmaceutical industries. EPS producing-lactic acid bacteria (LAB) have been reported to produce -fructans (both inulin and levan) as well as α-glucans through the action of sucrase enzymes fructansucrase (FS) and glucansucrase (GS). The Weissella confusa strain MBFCNC-2(1), isolated from a sample of a local beverage, and was shown to possess the open reading frame (ORF) of the FS coding gene (ftf) with high similarity to a putative inulosucrase of L. reuteri, which was designated ftfCNC-2(1). In total, 2,137 bp of full length ftfCNC-2(1) DNA was successfully cloned and sequenced, and a putative FTF protein of 900 amino acids was analyzed. A core region was identified as an active domain belonging to glycoside hydrolase 68. The N-terminal region contained a signal sequence followed by a sequence similar to the N-terminus of gtfKg15 (putatively identified as glucansucrase); a putative substrate binding domain was interestingly shown to be Serine Threonin rich. The C-terminus region contained an LPXTG motif, a cell wall anchoring domain, which was also Threonin rich. For further study, the ftfCNC-2(1) gene was successfully subcloned into a shuttle vector for future expression in either E. coli or B. subtilis.