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We studied the interaction of spectrin dimers from human erythrocytes with (bilayer and monolayer) model membranes of mixtures of dimyristoylphosphatidylethanolamine, dimyristoylphosphatidylcholine, and dimyristoylphosphatidylserine (DMPS) by densitometric evaluation of phase transitions and phase boundaries, film balance experiments, and microfluorescence. We demonstrate that spectrin readily adsorbs to mixed bilayers and monolayers even in the presence of small DMPS concentrations (30 mol %) whereas no appreciable interaction with lamellae containing zwitterionic lipids alone is observed. The selectivity of the DMPS/spectrin interaction is established by quantitative evaluation of the shifts of the phase boundaries (liquidus and solidus line) caused by the lipid/protein interaction as a function of the composition of the binary lipid mixtures. Quantitative information about the free energy of the lipid/protein interaction is obtained by computer simulation of the phase diagram of the lipid mixture in the absence or in the presence of a very small molar fraction of the protein and comparison of calculated and measured shifts. A binding energy of about 10(-17) J per spectrin molecule is found. The present perturbation method can be generalized to study selective lipid/protein interaction mechanisms in ternary or higher component mixtures. The present results provide evidence that in addition to the binding to band III, spectrin may also couple directly to the lipid moiety of the inner monolayer of erythrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Mixed vesicles of dimyristoylphosphatidylcholine (DMPC) and a polymerizable lipid containing one diene group per chain are studied by freeze fracture electron microscopy and by the photobleaching (fluorescence recovery after photobleaching) technique. Large thin-walled vesicles of some micron in diameter become more stable after photochemical polymerization. Before polymerization bilayers of the diene lipid exhibit a liquid crystal-to-gel transition at Tg = 31 degrees C. Upon polymerization the transition remains but shifts to a slightly higher temperature (Tg* = 34 degrees C). The transitions in both cases are accompanied by a freezing in of the lateral mobilities. The mixed vesicle exhibits lateral phase separation after polymerization. Before polymerization the two lipids appear miscible at all compositions in the fluid state and at DMPC concentrations at or below 50 mol % in the solid state. After polymerization a two-dimensional solution of the polymer in DMPC is obtained at T greater than Tg*, while lateral phase segregation into DMPC-rich domains and patches of the polymer is observed at T less than Tg*. The domain structure appears identical irrespective of whether polymerization is performed at T greater than Tg or at T less than Tg. A typical value of the diameter of the polymerized lipid domains (approximately 400 A) indicates a rather small aggregation number (N less than 100 monomers). The lateral diffusion coefficient in butadiene-lipid bilayers only decreases from D1 = 3.10(-7) cm2/s to D1 = 8.10(-8) cm2/s (that is by a factor of 4) upon polymerization. This is consistent with the freeze fracture finding of a small aggregation number. We point out the similarities of the mixed vesicles with plasma membranes coupled to the cytoskeleton.

Macromolecular adsorption is known to occur as a complex process, often in a series of steps. Several models are discussed in the literature which describe the microscopic structure of the adsorbate. In the present study we investigated the adsorption of hen egg white lysozyme on alkylated silicon oxide surfaces. A combination of fluorescence excitation in the evanescent field and fluorescence recovery after photobleaching allowed us to measure the amount of adsorbed fluorescent lysozyme and the equilibrium exchange kinetics with molecules in solution. We found that a model with at least three classes of adsorbed molecules is necessary to describe the experimental results. A first layer is formed by the molecules which adsorb within a short time after the beginning of the incubation. These molecules make up approximately 65% of the final coverage. They are quasi-irreversibly adsorbed and do not measurably exchange with bulk molecules within one day even at temperatures up to 55 degrees C. A second layer, which reaches equilibrium only after several hours of incubation, shows a pronounced exchange with bulk molecules. The on-off kinetics show a distinct temperature dependence from which an activation barrier of delta E approximately 22 kcal/mol is derived. A third layer of molecules that exchange rapidly with the bulk can be seen to comprise approximately 10% of the total coverage. The exchange rate is on the order of fractions of a second. The binding of the latter two classes of adsorbed molecules is exothermic. From the temperature dependence of the coverage, the binding enthalpy of the slowly exchanging layer was estimated to be delta Hads approximately 3.8 kcal/mol. The second and third class of molecules remain enzymatically active as a muramidase, which was tested by the lysis of the cell walls of Micrococcus lysodeiktikus. The molecules in the first layer, on the other hand, showed no enzymatic activity.

variations.41 Finally, it would be desirable to have experimental measurements of the and if, of Preston's polymer systems to further test the theory.Acknowledgment. This work was performed under the auspices of the U.S. Department of Energy by the Lawrence Livermore (41) Also suggested by Svensson-Rilbe, H., private communication.National Laboratory under contract W-7405-Eng-48. We are grateful to Dr. R. N. Schock and the LLNL Earth Sciences Division for support of V.V., to OBES Geosciences for support of D.G.M., and to Sue Frumenti for the word processing of the manuscript. We are also indebted to Dr.