Hydrolysis-transglycosylation of sucrose and production of β-(2→1)-fructan by inulosucrase from Neobacillus drentensis 57N

Abstract

Inulin, β-(2→1)-fructan, is a beneficial polysaccharide used as a functional food ingredient. Microbial inulosucrases (ISs), catalyzing β-(2→1)-transfructosylation, produce β-(2→1)-fructan from sucrose. In this study, we identified a new IS (NdIS) from the soil isolate, Neobacillus drentensis 57N. Sequence analysis revealed that, like other Bacillaceae ISs, NdIS consists of a glycoside hydrolase family 68 domain and shares most of the 1-kestose-binding residues of the archaeal IS, InuHj. Native and recombinant NdIS were characterized. NdIS is a homotetramer. It does not require calcium for activity. High performance liquid chromatography and 13C-nuclear magnetic resonance indicated that NdIS catalyzed the hydrolysis and β-(2→1)-transfructosylation of sucrose to synthesize β-(2→1)-fructan with chain lengths of 42 or more residues. The rate dependence on sucrose concentration followed hydrolysis-transglycosylation kinetics, and a 50% transglycosylation ratio was obtained at 344 m m sucrose. These results suggest that transfructosylation from sucrose to β-(2→1)-fructan occurs predominantly to elongate the fructan chain because sucrose is an unfavorable acceptor.

Keywords: Neobacillus drentensis 57N; glycoside hydrolase family 68; inulin; inulosucrase; sucrose.