Annular lipids determine the ATPase activity of a calcium transport protein complexed with dipalmitoyllecithin

Abstract

Pure complexes of dipalmitoyllecithin (DPL, 16:0, 16:0) with Ca2+, Mg2+ dependent ATPase from [rabbit skeletal muscle] sarcoplasmic reticulum are unusual in retaining significant ATPase activity down to about 30.degree. C, well below the transition temperature of the pure lipid at 41.degree. C. A minimum of about 35 lipid molecules/ATPase is required to maintain maximal ATPase activity, but the complexes are progressively and irreversibly inactivated at lower lipid to protein ratios. Complexes containing more than the minimum lipid requirement show very similar temperature profiles of activity above 30.degree. C over a wide range of lipid to protein ratios, up to 1500:1. Spin-label studies indicate that at lipid to protein ratios of less than about 30 lipids/ATPase, no DPL phase transition can be detected, but at all higher ratios a phase transition occurs at about 41.degree. C. In all of these complexes there are breaks in the Arrhenius plots of ATPase activity at 27-32.degree. C and at 37.5-38.5.degree. C. Experiments with perturbing agents such as cholesterol and benzyl alcohol, which have well-defined effects on the DPL phase transition, indicate that these breaks in the Arrhenius plots of ATPase activity cannot be attributed to a depressed and broadened phase transition in the lipids near the protein molecules. These results are interpreted as evidence for a phospholipid annulus of at least 30 lipid molecules which interact directly with the ATPase and cannot undergo a phase transition at 41.degree. C. This structural interaction of the ATPase with the annular DPL molecules has a predominant effect in determining the form of the temperature-activity profiles. However, the perturbation of the DPL phase transition does not extend significantly beyond the annulus, since a phase transition which starts at 41.degree. C can be detected as soon as extraannular lipid is present in the complexes. It may be a general feature of membrane structure that penetrant membrane proteins interact with their immediate lipid environment so as to cause only a minimal perturbation of the lipid bilayer.