Purification and characterization of heat-stable exo-inulinase from Streptomyces sp.

The extracellular exo-inulinase from Streptomyces sp. was purified from the culture broth by ammonium sulphate precipitation, followed by successive chromatographies on DEAE-Sephacel and ConA-CL Agarose columns. The enzyme was purified 18-folds with 4.8% activity yield from the starting culture broth. The purified enzyme gave a single band on gel electrophoresis and its molecular weight was estimated to be 45 kDa. The optimum temperature and pH for enzyme activity were 70 °C and 6.0, respectively. All metal salts except NaNO3, EDTA and HgCl2 were tolerated and did not adversely affect inulinase activity. The effect of thermal stabilizers on inulinase activity was studied. Consequently, glycerol and mannitol have been shown to have a protective effect on enzyme activity. As compared to sucrose and raffinose the purified enzyme had a lower Km (1.63 mM) and higher Vmax (450 mM) for inulin. All these conditions make Streptomyces sp., a potential candidate for industrial enzymatic production of high fructose syrup and in other large-scale biotechnological processes.