(Na+,K+)-ATPase of mammalian brain: effects of temperatures on cation and ATP interactions regulating phosphatase activity.

The effects of temperature on interactions between univalent cations or ATP and the p-nitrophenylphosphatase activity associated with brain (Na+,K+)-ATPase were examined. The apparent affinity for K+ activation under conditions favoring the moderate affinity site was temperature dependent, increasing with decreasing temperature. A comparison of univalent cations showed that the negative apparent delta H and delta S for cation binding increased with increasing apparent cation affinity. In contrast to the case with the moderate affinity sites, apparent affinity for the high affinity K+ site was independent of temperature. As temperature decreased, properties of moderate affinity site binding approached those of the high affinity site. The temperature dependence of ATP inhibition was opposite to that for K+ activation, with positive apparent delta H and delta S. The apparent delta H and delta S for cation binding approached those for the overall conformational change to K+-sensitive enzyme as cation affinity increased. These data suggest that E2, the K+-sensitive form of (Na+,K+)-ATPase, is stabilized by forces that require a decrease in entropy, explaining the predominant existence of E1 at physiologic temperatures. A conformational change leading to stabilization of E2 at higher temperatures can be produced by binding of univalent cations to a moderate affinity, presumably intracellular, site. This effect is counteracted by ATP. ATP also appears to alter the selectivity of this site to favor Na+ over K+ binding.