Die Reindarstellung der Kynureninase

Abstract

The enzyme catalyzes the hydrolytic cleavage of kynurenine to anthranilic acid and alanine. The object of the study was to determine whether the reaction is catalyzed by a single enzyme or by an enzyme complex. The starting material was hog liver acetone powder. Inactive protein was removed from an extract of this acetone powder by means of heat coagulation and ammonium sulfate precipitation. The enzyme itself was then precipitated by 52% saturation with ammonium sulfate and was dialyzed. It was then adsorbed to calcium phosphate, and fractionally eluted with phosphate buffer. The purest fraction was concentrated 150 times. It was homogeneous as judged by electrophoresis and ultracentrifugation. This preparation catalyzed the hydrolysis of kynurenine; kynureninase is thus taken to be a single enzyme. The enzyme was found to contain 0.012% pyridoxal phosphate; however, the true pyridoxal-phosphate content is most likely much higher, since the purified enzyme was activated by it, and some of it is therefore probably split off during the purification procedure. Metal ions had no activating effect.