Sepharose 4B 固定化チアミナーゼIの酵素学的性質

The bacterial thiaminases I produced by B. thiaminolyticus strain B and B.thiaminolyticus Matsukawa et Misawa(T.S.B. and T.M.M., respectively) were immobilized by CNBr-activated Sepharose 4B, and their enzymological properties were examined.
The total activities for the immobilized T.S.B. and T.M.M. obtained were 165 and 73 percents, respectively, of those for the native preparations. Although optimal pH and optimal temperature were virtually inchanged by the immobilization, pH- and thermostabilities were considerably increased.
The kinetic parameters(K_m and V_<max>), determined for the base-exchange reaction between thiamine and pyridine at pH 5.5 and 50℃, showed that the immobilization resulted in appreciable increases in both V_<max> and K_m values, except for the K_m value of a thiamine-T.M.M. system.
Upon being stored at 4℃, the immobilized T.M.M. was very stable and its activity was virtually unchanged at 315 days after immobilization whereas the immobilized T.S.B. was considerably unstable. The stability of the immobilized T.M.M. against continuous utilization, determined by a batch method, was also superior to that of the immobilized T.S.B.
These results suggest that the immobilized T.M.M. is more excellent than the immobilized T.S.B. as judged from the viewpoint of their use for catalysts.