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

摘要

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℃ and 6.0, respectively. All metal salts except NaNO_3, EDTA and HgCl_2 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 K_m (1.63 mM) and higher F_(max) (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 biotechnolog-ical processes.