Mg²⁺-Dependent Adenosine Triphosphatase in Isolated Synaptic Vesicles

Attribution of Most of the Enzymic Activity to an Actomyosin-Like Protein

Abstract

Synaptic vesicle fractions, which were isolated from bovine cerebral cortex, caudatolenticular nuclei, and chick brain according to Kadota and Kadota (1), contained an Mg²⁺-dependent ATPase showing maximum activity at around pH 6.5-6.8 in common. The ATPase was similar to that in cow-adrenal chromaffin granules as regards the optimum pH (16) and susceptibility to inhibition by NEM (18). SDS-polyacrylamide gel electrophoresis revealed that the vesicle fractions contained protein components which were tentatively identified as a myosin-like protein (220, 000 daltons), tubulin (54, 000), and actin (45, 000), as reported previously (6, 17, 19). Further purification of the vesicles by sucrose density gradient centrifugation brought about a decrease in the content of the myosin-like protein, accompanied by a decrease in the Mg²⁺-ATPase activity. These results suggest that most of the Mg²⁺-ATPase in the vesicle fractions is due to the (acto)myosin-like protein, whose Mg²⁺-ATPase shows maximum activity at around pH 6.8 (21), and that the protein could be separated from the synaptic vesicles.
The synaptic vesicle fraction of caudatolenticular nuclei bound tritiated dopamine in a saturable manner at pH 6.7 and 25°C, but the binding was not accelerated by ATP. The pH-dependency of the binding of dopamine did not resemble that of the Mg²⁺-ATPase activity. Hence the Mg²⁺-ATPase appeared not to be involved in the uptake of dopamine by the synaptic vesicles.