Studies on the Chemistry of the Living Bark of the Black Locust Tree in Relation to Frost. VI. Amylase and Phosphorylase Systems of the Bark Tissues

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Abstract
Amylase and phosphorylase enzyme systems have been detected in the living bark tissue of the black locust tree. The optimal temperature for the amylase system during a short incubation period (27 minutes), at pH 6.6, was greater than 55[degree] C. With a longer incubation time (64 min.) there was pronounced inactivation of the enzyme at 55[degree] C and maximum activity was observed at 50[degree] C. The activation energy for the amylase catalyzed reaction was approximately 12,000 calories per mole. The optimal pH range in citrate-phosphate buffer for the amylase system at 39 [degree]C was 5.3-6.0. The products of hydrolysis of starch by the locust tree amylase were maltose, glucose, and dextrins. Synthesis of polysaccharide fromglucose-1-phosphate by the locust tree phosphorylase was dependent on the presence of a small amt. of polysaccharide primer. The reaction was reversible and approached an equilibrium ratio of inorganic phosphate to glucose-1-phosphate which was independent of the concn. of starch. This ratio was lowest at the highest pH values. The optimal temp, for action of the locust tree phosphorylase during 45 minutes, at pH 6.6, was approximately 50[degree] C. The optimal pH in citrate buffer was approximately 6.3. The amylase system of the black locust was inhibited by I-KI reagent, p-chloromercuribenzoate, phenylmercuric chloride, and phenylmercuric nitrate. The phosphorylase of the black locust was inhibited by I-KI reagent, p-chloromercuribenzoate, phenylmercuric chloride, and phenylmercuric nitrate, but it was not inhibited by iodoacetate. The inhibitions of both enzymes by phenylmercuric chloride and phenylmercuric nitrate were readily reversed by cysteine. These studies indicate that both the amylase and phosphorylase systems of the black locust tree are dependent on the integrity of sulfhydryl groups. However, the essential sulfhydryl groups of the phosphorylase are not as chemically reactive as those of the amylase.