Purification and characterization of a novel transfructosylating enzyme from Bacillus macerans EG-6

Abstract

A novel transfructosylating enzyme, which produces fructo-oligosaccharides from sucrose from Bacillus macerans EG-6 were purified 63.5-fold by ammonium sulphate precipitation (20-60%), CM-Sepharose CL 6B and fast protein liquid chromatographies on Resource Q, Phenyl-Superose HR 5/5 and Mono S (Pharmacia, Uppsala, Sweden). The minimum molecular mass of the purified enzyme was 66 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme was stable at the pH range of 5.0-7.0 and had an optimum pH at 5.0. The optimum temperature for enzyme activity was at 50°C. The oligosaccharide compositions in reaction products were significantly different on using the enzyme obtained from each purification step. For example, crude enzyme unusually produced selectively GF5- and GF6-fructo-oligosaccharide whereas purified enzyme produced mainly 1-kestose (GF2) and nystose (GF3) as in the case of other transfructosylating enzymes.