Effect of the biochemical state of the Ca-ATPase protein of scallop sarcoplasmic reticulum on its interaction with trans-parinaric acid.
The polyene fluorescent probe trans-parinaric acid (tPA) was used to compare lipid-protein interactions in the scallop fragmented sarcoplasmic reticulum (FSR) between biochemical states where the Ca-ATPase molecules were arranged differently in the membrane and had different tertiary conformations. The state of the bulk lipid phase was examined over the temperature range -3 to +32 degrees C by exciting the tPA directly at 320 nm. The state of the system close to the Ca-ATPase protein was followed over the same temperature range by indirectly exciting the tPA through resonance energy transfer from the Ca-ATPase protein, with approximately one twenty-fifth the quantum yield of the directly excited probe. Raising the tPA/lipid ratio in the membrane to high levels (approx. 1:9), caused the quantum yield of indirectly excited tPA to reach a maximum, which may reflect saturation of the annular lipid phase with the probe, or contribution to the fluorescence from indirectly excited tPA bound directly to the protein. In the presence of 0.1 M KCl, a thermal perturbation was observed at approx. 7 degrees C using indirect excitation when the Ca(2+)-binding sites on the Ca-ATPase were occupied, and the subunits were disorganized. This transition was not detected in the presence of 0.1 M KCl and EGTA, when the Ca(2+)-binding sites were empty, and the Ca-ATPase subunits were organized in dimeric arrays. The transition seen with the E1(Ca2+)2 form of the membrane may involve an event at the protein/lipid interface, or a change in the environment of tPA bound to the Ca-ATPase. The temperature at which the perturbation occurs is close to that of a discontinuity in the Arrhenius plot of the Ca-ATPase enzyme activity determined in the presence of 0.1 M KCl (Kalabokis, V.N. and Hardwicke, P.M.D. (1988) J. Biol. Chem. 263, 15184-15188). No perturbation was observed in the bulk properties of the lipid component of the membrane in either the E1(Ca2+)2 or E2 states.
Kalabokis
V N
VN
Department of Medical Biochemistry, Southern Illinois University, Carbondale.
Hardwicke
P M
PM
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000737:chemistry* / Q000378:metabolism; D005231:Fatty Acids, Unsaturated / Q000378:metabolism*; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000378:metabolism*; D008974:Mollusca; D010743:Phospholipids / Q000378:metabolism; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013696:Temperature
D005231:Fatty Acids, Unsaturated; D005456:Fluorescent Dyes; D010743:Phospholipids; D000252:Calcium-Transporting ATPases; C013641:parinaric acid; D002118:Calcium