The Properties of ATPase of Synaptic Vesicle Fraction

Publisher Website
Google Scholar
Abstract
A synaptic vesicle fraction was prepared from the neocortex of guinea pig and the properties of its ATPase [EG 3.6.1.3] were investigated. The following results were obtained. 1. ATP is split at a rate of 4.5 μmoles/mg. protein/30 minutes at 20° C and pH7.4 in the presence of 4.0 mM Mg++, and the rate of splitting of ADP is about 20% of that of ATP. AMP does not serve as a substrate. 2. ATPase is activated about 10 fold by 4.0 mM Ca++ or 8.0 mM Mg++. The Km for Ca++ is 4.5 × 10−4M and that for Mg++ is 1.9 × 10−4M. 3. 1.0 mM EDTA abolishes ATPase activity completely while 1.0 mM GEDTA has only a slight inhibitory effect. 4.0 mM Mg++ and Ca++ reverse the inhibitions caused by these chelating reagents. 4. In the absence of divalent cation, 0.10 M K+ stimulates ATPase activity, while 0.60 M K+ is inhibitory. The effect of Na+ is similar to that of K+.1.0 mM EDTA abolishes the effect of Na+or K+on ATPase activity, whereas in the presence of 1.0 mM GEDTA the effect of Na+ or K+ is similar to that in the presence of Mg++. When Mg++ is present, the activating effect of 0.10M Na+ or K+ is not clear, but the inhibitory effect 0.60 M Na+ or K+ increases. 5.Thiol blocking reagents depress ATPase activity.Glutathione and cysteine reverse the inhibition caused by these thiol blocking reagents. 6.Treatment by phospholipase C [EC 3.1.4.3], freezing and thawing, or 10 mM desoxycholic acid depress ATPase activity. 7.There is a slight competition between Mg++ and Ca++. 5.0 × 10−4M DNP has no effect. Oligomycin and atractylate do not decrease the activation induced by Mg++. No reduction of cytochrome c is seen with 50 mM succinic acid as substrate. 8. Na+plus K+ stimulates ATPase slighdy in the presence of Mg++ and ouabain abolishes this effect. With a more purified synaptic vesicle preparation, however, the inhibitory effect of is incomplete.