PHOSPHOLAMBAN-DEPENDENT EFFECTS OF C(12)E(18) ON CALCIUM-TRANSPORT AND MOLECULAR-DYNAMICS IN CARDIAC SARCOPLASMIC-RETICULUM

We have studied the effects of the nonionic detergent C(12)E(8) On Ca- ATPase enzymatic activity and oligomeric state (detected by time-resol ved phosphorescence anisotropy, TPA) in skeletal and cardiac sarcoplas mic reticulum (SR). In skeletal SR, C(12)E(8) inhibits the Ca-ATPase, both at high (micromolar and above) and low (submicromolar) Ca. In car diac SR, C(12)E(8) inhibits at high Ca but activates at low Ca. Thus C (12)E(8) activates enzymatic activity only in cardiac SR and only unde r conditions (submicromolar Ca) where phospholamban (PLB) regulates (i nhibits) the enzyme [Lu, Y.-Z., & Kirchberger, M. A. (1994) Biochemist ry 33, 5056-5062]. TPA of skeletal SR at low and high Ca demonstrates that C(12)E(8) induces aggregation of ATPase monomers and small oligom ers. C(12)E(8) also aggregates the Ca-ATPase in cardiac SR at high Ca. In cardiac SR at low Ca, the Ca-ATPase is already highly aggregated, and C(12)E(8) partially dissociates these aggregates. Thus the TPA res ults provide a simple physical explanation for the functional effects: C(12)E(8) inhibits the ATPase when it aggregates the enzyme (skeletal SR at high and low Ca; cardiac SR at high Ca), and the detergent acti vates when it dissociates ATPase oligomers (cardiac SR at low Ca). C(1 2)E(8) stabilizes the E2P conformation of the Ca-ATPase with respect t o the E2 conformation, and this stabilization is PLB-dependent. Both t he physical and the functional effects of C(12)E(8) On the Ca-ATPase a re PLB-dependent, with C(12)E(8) reversing the effects of PLB. The res ults provide insight into the mechanism by which PLB regulates the Ca- ATPase in cardiac SR.