Purification and characterization of fructan and fructansucrase from Lactobacillus fermentum AKJ15 isolated from Kodo ko jaanr, a fermented beverage from north-eastern Himalayas

Fructansucrase and fructan produced from Lactobacillus fermentum AKJ15 were isolated from seeds of Kodo ko jaanr, a fermented mild-alcoholic beverage prepared in North East India. The strain was identified by 16S rRNA gene sequence analysis and biochemical characterization. The strain displayed maximum fructansucrase activity of 4.3 U/ml (1.02 U/mg) at 28 degrees C at 180 rpm. The enzyme purified by polyethylene glycol-400 gave specific activity of 5 U/mg and showed 90 kDa band on non-denaturing Sodium Dodecyl Sulphate-Poly Acrylamide Gel Electrophoresis (SDS-PAGE). The purified enzyme confirmed the presence of fructan by periodic acid Schiff's staining which showed magenta colour bands with both sucrose and raffinose. The strain produced 10.2 mg/ml fructan in broth under optimized culture conditions. The purified fructansucrase displayed V-max of 5.42 U/mg and K-m of 16.65 mM. The enzyme showed maximum activity at 30 degrees C and at pH 5. The structure of fructan was analysed by H-1 and C-13 NMR spectra confirming beta-(2-1) and beta-(2-6) linkages.