Data are presented that lead to an alternative model for the organization and molecular dynamics of lipid molecules near the Ca2+-stimulated, Mg2+-dependent adenosinetriphosphatase (Ca2+-ATPase; ATP phosphohydrolase, EC 3.6.1.3) of sarcoplasmic reticulum. Measurements of the steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene in progressively delipidated sarcoplasmic reticulum membranes have been quantitatively interpreted in terms of a layer of lipid of high anisotropy (the lipid annulus) coexisting with lipid layers of very low anisotropy. In addition, the Ca2+-ATPase has been reconstituted into pure 1,2-dipentadecanoyl 3-sn-phosphatidylcholine membranes over a range of lipid-to-protein ratios. High-sensitivity differential scanning calorimetry has demonstrated that roughly 30 lipid molecules per Ca2+-ATPase molecule (annular lipids) fail to undergo a calorimetrically detectable phase transition in the temperature range 4-44 degrees C. Roughly 100 lipid molecules beyond the annulus undergo a detectable phase transition at a temperature below the phase transition of pure lipid and with an enthalpy change [4.2 kcal/mol (1 kcal = 4.18 kJ)] about half that observed for pure lipid vesicles (7.7-7.8 kcal/mol). We propose that both the fluorometric and calorimetric data are consistent with a model in which a motionally inhibited lipid annulus is surrounded by a more extensive region of disrupted lipid packing order, which we have called the secondary lipid domain
