Effects of phospholipid acyl chain fluidity, phase transitions, and cholesterol on (Na+ + K+)-stimulated adenosine triphosphatase.

A soluble delipidized (Na + K)-stimulated ATPase was obtained from rabbit kidney outer medulla by the use of sodium deoxycholate. The activity of this enzyme was stimulated ZO-fold to an activity of 100 to 120 pmoles of Pi per mg of protein per hour by either phosphatidylserine or phosphatidylglycerol. The effect of the fluidity of the hydrocarbon region of the phospholipid bilayers on this re-activation of (Na + K)ATPase was systematically examined. Fatty acyl chains of known length and varying saturation were substituted on the phosphatidylglycerol moiety and the influence of temperature on the degree of re-activation was investigated. Discontinuities in Arrhenius plots of (Na + K)-ATPase activity were found and compared with the gel-to-liquidcrystalline transitions of these substituted phosphatidylglycerols as measured in a differential scanning calorimeter. The temperature at which the Arrhenius plot discontinuities occurred were from lo to 8” lower than the initial rise of the main endothermic peak for the series: dimyristoyl-, dipalmitoyl-, and distearoylphosphatidylglycerols. Dioleoylphosphatidylglycerol which does not undergo a phase transition within the experimental temperature range was found to activate at much lower concentrations than the saturated phosphatidylglycerols, and the Arrhenius plot did not show discontinuities. In-contrast, bovine brain phosphatidylserine with a heterogeneous fatty acyl chain composition showed an Arrhenius plot discontinuity which was 2” higher than the midpoint of the broad endothermic peak. The relation between the discontinuities in the Arrhenius plots of the enzyme activity and the lipid phase transitions is discussed. It is suggested that some of the discontinuities represent the completion of melting (upper limit for a broad endothermic transition), whereas others represent the beginning of melting (lower limit). For completely liquid membranes, the slopes of the Arrhenius plots are parallel and give an activation energy of approximately 15 Cal per mole.