OCCURRENCE OF 2 FORMS OF EXTRACELLULAR ENDOINULINASE FROM ASPERGILLUS-NIGER MUTANT-817

Aspergillus niger mutant 817 was generated from A. niger 12, a strain that produces high levels of inulinases irrespective of the carbon source, by gamma-irradiation with Co-60. A. niger 817 showed 4-fold higher inulinase activity than the wild-type strain 12 in the filtrate of a submerged culture with fructose. An extremely high ratio of inulinase to invertase activity (I/S ratio) of 880 was found in the culture filtrate. Two inulinases, P-IA and P-IB, were purified from the culture filtrate of A. niger 817 grown on sucrose by DEAE-Cellulofine A-500 and Q-Sepharose HP chromatographies. The enzymes were homogeneous as judged by SDS-polyacrylamide gel electrophoresis with apparent molecular weights of 70,000 for P-IA and 68,000 for P-IB. The specific activities were 350 U/mg for P-IA and 340 U/mg for P-IB, and about 3.5-fold higher than that reported for the wild-type endoinulinase P-III, The enzymes were active only toward inulin, and lacked activity toward sucrose, raffinose or levan. The main products of inulin hydrolysis were inulotriose and -tetraose. Thus, both inulinases exhibited the endo-type mode of action on inulin. The apparent K, values for inulin were 0.48 mM for P-IA and 0.50 mM for P-IB at 40 degrees C and pH 5.0, and lower than the 1.25 mM reported for the wild-type endoinulinase P-III.