Enzymatic production of glucosamine and chitooligosaccharides using newly isolated exo-β-d-glucosaminidase having transglycosylation activity

Exochitosanase secreting fungus (A. fumigatus IIT-004) was isolated from fish waste using 1 % (w/v) chitosan as sole carbon source after multistage screening. Chitosan-dependent exochitosanase enzyme production (6 IU ml-1) in log phase of growth (chitosan utilization rate 0.11 g g-1 cell h-1) was observed for Aspergillus fumigatus in chitosan minimal salt medium and there was no enzyme production in glucose medium. Enzyme production was found to be extracellular and subjected to purification by a number of steps like acetone fractionation as well as column chromatography. 40 % yield and 26-fold of enzyme purification was achieved after all the steps. Purified enzyme was characterized for optimum temperature, pH, ionic strength and substrate specificity. The K m and V max for purified exochitosanase enzyme was calculated to be 8 mg ml-1 and 5.2 × 10-6 mol mg-1 min-1. Enzyme was immobilized on polyacrylonitrile nanofibres membrane matrix by adsorption as well as amidination. Enzymatic production of glucosamine was achieved using various chitosan substrates by free/immobilized exochitosanase and compared. Isolated and purified exochitosanase also showed transglycosylation activity.