Temperature dependence of rotational dynamics of protein and lipid in sarcoplasmic reticulum membranes.
We have investigated the relationship between function and molecular dynamics of both the lipid and the Ca-ATPase protein in sarcoplasmic reticulum (SR), using temperature as a means of altering both activity and rotational dynamics. Conventional and saturation-transfer electron paramagnetic resonance (EPR) was used to probe rotational motions of spin-labels attached either to fatty acid hydrocarbon chains or to the Ca-ATPase sulfhydryl groups in SR. EPR studies were also performed on aqueous dispersions of extracted SR lipids, in order to study intrinsic lipid properties independent of the protein. While an Arrhenius plot of the Ca-ATPase activity exhibits a clear change in slope at 20 degrees C, Arrhenius plots of lipid hydrocarbon chain mobility are linear, indicating that an abrupt thermotropic change in the lipid hydrocarbon phase is not responsible for the Arrhenius break in enzymatic activity. The presence of protein was found to decrease the average hydrocarbon chain mobility, but linear Arrhenius plots were observed both in the intact SR and in extracted lipids. Lipid EPR spectra were analyzed by procedures that prevent the production of artifactual breaks in the Arrhenius plots. Similarly, using sample preparations and spectral analysis methods that minimize the temperature-dependent contribution of local probe mobility to the spectra of spin-labeled Ca-ATPase, we find that Arrhenius plots of overall protein rotational mobility also exhibit no change in slope. The activation energy for protein mobility is the same as that of ATPase activity above 20 degrees C; we discuss the possibility that overall protein mobility may be essential to the rate-limiting step above 20 degrees C.
D000818:Animals; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D005227:Fatty Acids / Q000032:analysis; D007700:Kinetics; D008563:Membrane Lipids / Q000032:analysis; D008565:Membrane Proteins / Q000378:metabolism*; D009132:Muscles / Q000201:enzymology; D011817:Rabbits; D012399:Rotation; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013113:Spin Labels; D013816:Thermodynamics
D005227:Fatty Acids; D008563:Membrane Lipids; D008565:Membrane Proteins; D013113:Spin Labels; D000252:Calcium-Transporting ATPases