| Abstract: | Solubilization of the ATPase of sarcoplasmic reticulum vesicles with the nonionic detergent dodecyl non-aoxyethylene alcohol (C12E9) resulted in a large (about 5-fold) increase in its Ca2+ ATPase activity. Measurements using a calcium ionophore suggest this activation was the result of rendering the vesicles permeable to calcium. Complete activity is preserved at a detergent concentration range in which the detergent is complexed with the monomeric form of the ATPase, as measured by Sepharose 6B chromatography. Using a calibrated column, we found the C12E9 complex to have a Stokes radius of 55 A. As measured by time-resolved fluorescence anisotropy decay experiments, it had a rotational correlation coefficient of 214 ns, which is equivalent to a Stokes radius of 59 A. The axial ratio of the corresponding ellipsoid of revolution is calculated to be 5 to 6, indicating the complex is quite asymmetric. Like the vesicular form of the ATPase, the detergent-solubilized monomeric form bound with high affinity about 9 nmol of Ca2+/mg of protein. Also, like the vesicular enzyme, the solubilized form displayed a Ca2+ dependence of the activation of ATP hydrolysis which was cooperative (Hill coefficient 1.8). These results suggest that the calcium sites interact intramolecularly. |
