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Abstract

The terminal phosphate group of ATP was transferred to ADP by an enzyme present in the soluble core proteins of adrenal medulla catecholamine storage vesicles. It was purified 10-30-fold by DEAE Sephadex chromatography (Fraction I). The enzyme required divalent metal ions for activation; Mn2+ was almost as effective as Mg2+, but Ca2+ was only a weak activator. Activation by Mg2+ took place over a very narrow concentration range (0.5-3 mM). The specificity of the enzyme activity to nucleoside triphosphates was broad, to the nucleoside diphosphates narrow, favouring adenosine diphosphate. In dependence on the pH the activity increased from pH 4 to pH 7 and remained constantly high between pH 7 and 9. The Arrhenius plot was linear between 5 and 70 degrees C, with an activation energy of 11.1 kcal/mol. The phosphoryl group transfer reaction depended on the function of thiol groups; p-hydroxymercuribenzoate inhibited 50% of the enzyme activity; dithioerythritol reactivated it completely. Gel electrophoresis revealed that in Fraction I, a protein of molecular weight about 45,000, was enriched compared with the total proteins. The enzyme-enriched Fraction I differed significantly in its relative amino acid composition from that of the total soluble proteins; in general, the acidic amino acids were reduced and the more basic acids enhanced.