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In fluid monolayers approaching collapse, phospholipids and their complexes with diacylglycerols hinder adsorption to the monolayer of the amphipathic protein, colipase. Herein, a statistical, free-area model, analogous to that used to analyze two-dimensional lipid diffusion, is developed to describe regulation by lipids of the initial rate of protein adsorption from the bulk aqueous phase to the lipid-water interface. It is successfully applied to rate data for colipase adsorption to phospholipid alone and yields realistic values of the two model parameters; the phospholipid excluded area and the critical free surface area required to initiate adsorption. The model is further developed and applied to analyze colipase adsorption rates to mixed monolayers of phospholipid and phospholipid-diacylglycerol complexes. The results are consistent with complexes being stably associated over the physiologically relevant range of lipid packing densities and being randomly distributed with uncomplexed phospholipid molecules. Thus, complexes should form in fluid regions of cellular membranes at sites of diacylglycerol generation. If so, by analogy with the behavior of colipase, increasing diacylglycerol may not trigger translocation of some amphipathic peripheral proteins until its abundance locally exceeds its mole fraction in complexes with membrane phospholipids.

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Lipases are extracellular peripheral proteins that act at the surface of lipid emulsions stabilized, typically, by phospholipids. At a critical composition lipase activity toward substrates in phospholipid monolayers is discontinuously switched on by a small increase in substrate mole fraction. This occurs in part because lipase binding is inhibited by phospholipids. Binding of the lipase cofactor, colipase, is also inhibited by phospholipids. The initial rate of colipase binding increases abruptly at a substrate mole fraction that is approximately half the critical composition for lipase activity and just above that in substrate-phospholipid complexes. Moreover, complex collapse areas show an approximately 1:1 correlation with phospholipid excluded areas determined from an analysis of colipase adsorption rates. Thus, complexes inhibit colipase binding rate. Additionally, the switching of lipase activity likely occurs when uncomplexed substrate becomes the majority species in the interface. Lipase substrates, e.g. diacylglycerols, are typically the same lipids generated in the cytoplasmic surface of the plasma membrane of stimulated cells. As colipase binding is nonspecific and complexes involving lipase substrates form on the basis of lipid-lipid interactions alone, complexes should form in the plasma membrane of stimulated cells and may regulate protein translocation to the membrane.

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Colipase, a cofactor of pancreatic triacylglycerol lipase, binds to surfaces of lipolysis reactants, like fatty acid and diacylglycerol, but not to the nonsubstrate phosphatidylcholine. The initial rate of colipase binding to fluid, single-phase lipid monolayers was used to characterize the interfacial requirements for its adsorption. Colipase adsorption rates to phosphatidylcholine/reactant mixed monolayers depended strongly on lipid composition and packing. Paradoxically, reactants lowered colipase adsorption rates only if phosphatidylcholine was present. This suggests that interactions between phosphatidylcholine and reactants create dynamic complexes that impede colipase adsorption. Complex formation was independently verified by physical measurements. Colipase binding rate depends nonlinearly on the two-dimensional concentration of phosphatidylcholine. This suggests that binding is initiated by a cluster of nonexcluded surface sites smaller than the area occupied by a bound colipase. Binding rates are mathematically consistent with this mechanism. Moreover, for each phosphatidylcholine-reactant pair, the complex area obtained from the analysis of binding rates agrees well with the independently measured collapse area of the complex. The dynamic complexes between phosphatidylcholine and lipids, like diacylglycerols, exist independently of the presence of colipase. Thus, our results suggest that lipid complexes may regulate the fluxes of other proteins to membranes during, for example, lipid-mediated signaling events in cells.

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In this paper the geometrical properties of gel and fluid clusters of equimolar dimyristoylphosphatidylcholine/distearoylphosphatidylcholine (DMPC/DSPC) lipid bilayers are calculated by using an Ising-type model (Sugar, I. P., T. E. Thompson, and R. L. Biltonen. 1999. Biophys. J. 76:2099-2110). The model is able to predict the following properties in agreement with the respective experimental data: the excess heat capacity curves, fluorescence recovery after photobleaching (FRAP) threshold temperatures at different mixing ratios, the most frequent center-to-center distance between DSPC clusters, and the fractal dimension of gel clusters. In agreement with the neutron diffraction and fluorescence microscopy data, the simulations show that below the percolation threshold temperature of gel clusters many nanometer-size gel clusters co-exist with one large gel cluster of size comparable with the membrane surface area. With increasing temperature the calculated effective fractal dimension and capacity dimension of gel and fluid clusters decrease and increase, respectively, within the (0, 2) interval. In the region of the gel-to-fluid transition the following geometrical properties are independent from the temperature and the state of the cluster: 1) the cluster perimeter linearly increases with the number of cluster arms at a rate of 8.2 nm/arm; 2) the average number of inner islands in a cluster increases with increasing cluster size, S, according to a power function of 0.00427 x S(1.3); 3) the following exponential function describes the average size of an inner island versus the size of the host cluster, S: 1 + 1.09(1 - e(-0.0072xS)). By means of the equations describing the average geometry of the clusters the process of the association of clusters is investigated.

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In this paper, we describe a relatively simple lattice model of a two-component, two-state phospholipid bilayer. Application of Monte Carlo methods to this model permits simulation of the observed excess heat capacity versus temperature curves of dimyristoylphosphatidylcholine (DMPC)/distearoylphosphatidylcholine (DSPC) mixtures as well as the lateral distributions of the components and properties related to these distributions. The analysis of the bilayer energy distribution functions reveals that the gel-fluid transition is a continuous transition for DMPC, DSPC, and all DMPC/DSPC mixtures. A comparison of the thermodynamic properties of DMPC/DSPC mixtures with the configurational properties shows that the temperatures characteristics of the configurational properties correlate well with the maxima in the excess heat capacity curves rather than with the onset and completion temperatures of the gel-fluid transition. In the gel-fluid coexistence region, we also found excellent agreement between the threshold temperatures at different system compositions detected in fluorescence recovery after photobleaching experiments and the temperatures at which the percolation probability of the gel clusters is 0.36. At every composition, the calculated mole fraction of gel state molecules at the fluorescence recovery after photobleaching threshold is 0.34 and, at the percolation threshold of gel clusters, it is 0.24. The percolation threshold mole fraction of gel or fluid lipid depends on the packing geometry of the molecules and the interchain interactions. However, it is independent of temperature, system composition, and state of the percolating cluster.

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Nystatin isolated from Streptomyces is a polyene antibiotic that is frequently used in the treatment and prophylaxis of fungal infections. Here, the fractional sterol concentration dependencies of the partition coefficient for partitioning of nystatin into ergosterol/dimyristoyl-L-alpha-phosphatidylcholine (DMPC), cholesterol/DMPC, ergosterol/1-palmitoyl-2-oleoyl-L-alpha-phosphatidylcholine (POPC), and ergosterol/POPC/1-palmitoyl-2-oleoyl-L-alpha-phosphatidylethano lam ine (POPE) multilamellar vesicles have been determined fluorometrically at 37 degrees C using approximately 0.3-1.0 mol % sterol concentration increments over a wide concentration range (e.g., 18-54 mol % sterol). This unconventional approach of varying membrane sterol content, in contrast to previous studies using large sterol concentration increments (e.g., 10 mol %), leads to a striking observation. The partition coefficient of nystatin changes dramatically with membrane sterol content in a well-defined alternating manner, displaying a local minimum at or very close to the critical sterol mole fractions (e.g., 20.0, 22.2, 25.0, 33.3, 40.0, and 50.0 mol % sterol) predicted for sterols regularly distributed in either hexagonal or centered rectangular superlattices. In ergosterol/DMPC bilayers, for example, there is a >3-fold increase in nystatin partitioning with a minute change (approximately 1 mol %) in sterol content on either side of the critical sterol mole fraction, 25.0 mol %. These results provide semifunctional evidence supporting the sterol regular distribution model [Chong, P. L.-G. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 10069-10073]. More importantly, these results reveal a new membrane phenomenon, that is, that nystatin partitioning is affected by the extent of sterol regular distribution in the plane of the membrane. This phenomenon occurs not only in saturated (e.g., DMPC) but also in unsaturated (e.g., POPC) lipid membranes, and persists in the presence of polar headgroup heterogeneity (e.g., POPC/POPE). This membrane property points to a new method for studying the interactions of polyene antibiotics with sterol-containing membranes, and the need to consider the membrane sterol content of the target cells when administering nystatin or other polyene antibiotics.

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We have examined the fractional sterol concentration dependence of dehydroergosterol (DHE) fluorescence in DHE/cholesterol/dimyristoyl-L-alpha-phosphatidylcholine (DMPC), DHE/ergosterol/DMPC and DHE/cholesterol/dipalmitoyl-L-alpha-phosphatidylcholine (DPPC) liquid-crystalline bilayers. Fluorescence intensity and lifetime exhibit local minima (dips) whenever the total sterol mole fraction, irrespective of the DHE content, is near the critical mole fractions predicted for sterols being regularly distributed in hexagonal superlattices. This result provides evidence that all three of these naturally occurring sterols (e.g., cholesterol, ergosterol, and DHE) can be regularly distributed in the membrane and that the bulky tetracyclic ring of the sterols is the cause of regular distribution. Moreover, at the critical sterol mole fractions, the steady-state anisotropy of DHE fluorescence and the calculated rotational relaxation times exhibit distinct peaks, suggesting that membrane free volume reaches a local minimum at critical sterol mole fractions. This, combined with the well-known sterol condensing effect on lipid acyl chains, provides a new understanding of how variations in membrane sterol content change membrane free volume. In addition to the fluorescence dips/peaks corresponding to hexagonal superlattices, we have observed intermediate fluorescence dips/peaks at concentrations predicted by the centered rectangular superlattice model. However, the 22.2 mol% dip for centered rectangular superlattices in DHE/ergosterol/DMPC mixtures becomes diminished after long incubation (4 weeks), whereas on the same time frame the 22.2 mol% dip in DHE/cholesterol/DMPC mixtures remains discernible, suggesting that although all three of these sterols can be regularly distributed, subtle differences in sterol structure cause changes in lateral sterol organization in the membrane.

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Activation of beta-2 adrenoceptors (BAR) in smooth muscle preparations is associated with a rapid, reversible and incomplete receptor desensitization, resulting in a steady-state relaxation response to BAR agonists. Based on results from cell culture studies, we hypothesize that, in the isolated guinea pig trachea, this steady state is a result of a concurrent resensitization of desensitizing BAR. In tracheal segments maintained at mechanical tone (4-6 g), isoproterenol (ISO) and the partial BAR agonist salbutamol (SALB) elicited a monotonic, rapid (1-3 min) and reproducible relaxation response that could be maintained for up to 45 min and was completely reversed by propranolol. Similarly, tissues preconstricted with 0.1 microM carbachol (CARB) responded with a sustained relaxation response to ISO. In contrast, in tissues preconstricted with 0.3 to 10 microM CARB or with 75 mM KCl, the relaxation elicited by ISO was followed by a slow (20-30 min) and partial restoration of muscle tone ("fade"). The relaxation and fade were observed when CARB-constricted tissues were relaxed with SALB (0.2 or 10 microM) or 10 microM salmeterol. No response to SALB was observed when tissues were preconstricted with KCl. The fade met criteria for its classification as a homologous desensitization of the relaxation response at the BAR level. In desensitized washed tissues, a complete recovery of the original relaxation response could be detected within 60 min of drug removal. A propranolol- and ICI 118-551-sensitive steady state was achieved 30 to 35 min after the addition of BAR agonists to the isolated tissues. A three-compartment phenomenological kinetic model accurately described the observed data, defining one steady-state and three rate constants, describing relaxation (k1), desensitization (k2) and resensitization (k3). The values of k2 and k3 for the response to SALB and to salmeterol were significantly larger than those observed for ISO. In the presence of KCl, the values of k2 and k3 for the response to ISO were indistinguishable from those measured in the presence of CARB. Given the parameters defined by our model, we propose that desensitization and resensitization of BAR in the isolated guinea pig trachea are distinct concurrent processes whose net result actively maintains a sustained partial relaxation response to ISO, SALB or salmeterol. The component of resensitization in the presence of agonist may account for the clinical efficacy of inhaled BAR agonists.

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Our previous studies indicated that sterols (including cholesterol and dehydroergosterol) can be regularly distributed into hexagonal superlattices in the plane of liquid-crystalline phosphatidylcholine bilayers. It was suggested that regular and irregular regions coexist in the membrane. In the present study, we report supporting evidence for our sterol regular distribution model. We have examined the fractional concentration dependencies of dehydroergosterol (a naturally occurring cholesterol analogue) fluorescence intensity and lifetime in various phosphatidylcholine and sphingomyelin bilayers. Fluorescence intensity and lifetime dips have been observed at specific sterol mole fractions. At those mole fractions, the acrylamide quenching rate constant of dehydroergosterol fluorescence reaches a local maximum. Those mole fractions match the critical sterol mole fractions at which sterol molecules are expected to be regularly distributed into hexagonal superlattices. The results support the idea that the sterols in the regular region are embedded in the bilayer less deep than those in the irregular regions. We have also examined the fractional cholesterol concentration dependencies of diphenylhexatriene (DPH) fluorescence intensity, lifetime, and polarization in DMPC vesicles. DPH fluorescence intensity and polarization also exhibit distinct dips and peaks, respectively, at critical sterol mole fractions for hexagonal superlattices. However, DPH lifetime changes little with sterol mole fraction. As a comparison, the fluorescence properties of DHE and DPH behave differently in response to the formation of sterol regular distribution. Furthermore, finding evidence for sterol regular distribution in both phosphatidylcholine and sphingomyelin membranes raises the possibility that sterol regular distribution may occur within phospholipid/cholesterol enriched domains of real biological membranes.

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A new method for structure refinement, based on optimization of the variable target function (Sugár and Xu, Prog. Biophys. Mol. Biol. 58, 61, 1992), is examined in order to explore the conditions of high-resolution polypeptide structure determination. The method of variable target function was used to analyze simulated NOESY spectra of different spectral resolutions, of different signal/noise ratios, and of different number of mixing times. The goodness of the results was assessed by comparing the known polypeptide conformation with the optimized ones, as well as by comparing the simulated experimental spectra with the spectra back-calculated from the optimized structures. According to the test calculations, in order to get high-resolution polypeptide structures, the number of constraints should be larger than the number of dihedral angles, and the constraints should be distributed over the dihedral angles more or less homogeneously. The success rate of the variable-target-function method in finding high-resolution structures was high until a critical signal/noise ratio, while below this threshold the success rate was low. The critical signal/noise ratio could be decreased, however, by increasing the number of mixing times. The test calculations show that the variable-target-function method is capable of high-resolution polypeptide structure determination with a success rate of 85-90% under common experimental conditions, such as 10 constraints/residue, 40% noise/signal ratio, and six mixing times.

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A global optimization method for intensity-restrained structure refinement, based on variable target function (VTF) analysis, is illustrated using experimental data on a model peptide, gramicidin-S (GS) dissolved in DMSO. The method (referred to as VARTIGO for variable target intensity-restrained global optimization) involves minimization of a target function in which the range of NOE contacts is gradually increased in successive cycles of optimization in dihedral angle space. Several different starting conformations (including all-trans) have been tested to establish the validity of the method. Not all optimizations were successful, but these were readily identifiable from their large NOE R-factors. We also show that it is possible to simultaneously optimize the rotational correlation time along with the dihedral angles. The structural features of GS thus obtained from the successful optimizations are in excellent agreement with the available experimental data. A comparison is made with structures generated from an intensity-restrained single target function (STF) analysis. The results on GS suggest that VARTIGO refinement is capable of yielding better quality structures. Our work also underscores the need for a simultaneous analysis of different NOE R-factors in judging the quality of optimized structures. The NOESY data on GS in DMSO appear to provide evidence for the presence of two orientations for the ornithine side chain, in fast exchange. The NOESY spectra for this case were analyzed using a relaxation rate matrix which is a weighted average of the relaxation rate matrices for the individual conformations.

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The intramolecular dynamics of the excimer forming dipyrenyl lipids (DipynPC) of different chain lengths (n) in ethanol and in dimyristoylphosphatidycholine (DMPC) membranes was investigated by the use of frequency-domain fluorescence intensity decay technique. Based on a 3-state model, the extent of aggregation and rotational rate of the two intralipid pyrene moieties in the dipyrenyl lipids were estimated from the frequency-domain data. In ethanol (20 degrees C), the rotational rate for DipynPC increased progressively as n was varied from 4 to 12. At the gel (L beta)-to-liquid crystalline (L alpha) phase transition of DMPC (approximately 23 degrees C), the rotational rate increased and aggregation decreased significantly for Dipy10PC, whereas only the rotational rate was changed for Dipy4PC. In the presence of 30 mol% cholesterol, significant increases in both the rotational rate and aggregation were observed for Dipy10PC in both L beta and L alpha phases. However, for the case of Dipy4PC, an increase in the rotational rate but a decrease in the aggregation were noticed only in the L beta phase, and no similar changes were detected in the L alpha phase. Our results indicate differential effects of cholesterol on the conformational dynamics of acyl chains at different depths of the membranes.

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The intramolecular dynamics of the excimer-forming dipyrenyl lipids (DipynPE) of different chain lengths (n) in fully hydrated dioleoylphosphatidylethanolamine (DOPE) and dioleoylphosphatidylcholine (DOPC) binary mixtures was investigated by the use of frequency-domain fluorescence intensity dcay technique. Using a 3-state model (see companion paper), the extent of aggregation and rotational rate of the two covalently attached pyrene moieties in DipynPE were estimated from the frequency-domain data. At 1 degrees C, the rotational rate and aggregation for Dipy4PE and Dipy10PE were insensitive to DOPE% of the lipid bilayer. At 27 degrees C, the rotational rate decreased, whereas the aggregation increased steadily for Dipy10PE as the DOPE% of the bilayer increased from 0 to 80. However, an abrupt increase in the rotational rate and a decrease in the aggregation for Dipy10PE were detected as the DOPE% reached 100, at which point the membranes are in the inverted hexagonal (HII) phase. No similar changes were found for Dipy4PE. These results indicate that the presence of PE with large intrinsic-curvature increases the lateral stress at the region near the center of the bilayer, and that this stress can be relieved as the membranes enter the highly curved HII phase.

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In a previous study, we observed a series of dips in the plot of E/M (the ratio of excimer to monomer fluorescence intensity) versus the mole fraction of 1-palmitoyl-2-(10-pyrenyl)decanoyl-sn-glycerol-3-phosphatidylcholine (Pyr-PC) in Pyr-PC/DMPC binary mixtures at 30 degrees C. In the present study, we have characterized the physical nature of E/M dips in Pyr-PC/DMPC binary mixtures by varying pressure, temperature, and vesicle diameter. The E/M dips at 66.7 and at 71.4 mol% PyrPC in DMPC multilamellar vesicles remain discernible at 30-43 degrees C. At higher temperatures (e.g., 53 degrees C), the depth of the dip abruptly becomes smaller. This result agrees with the idea that E/M dips appear as a result of regular distribution of pyrene-labeled acyl chains into hexagonal super-lattices at critical mole fractions. Regular distribution is a self-ordering phenomenon. Usually, in self-ordered systems, the number of structural defects increases with increasing temperature, and thermal fluctuations eventually result in an order-to-disorder transition. The effect of vesicle diameter on the E/M dip at 66.7 mol% Pyr-PC in DMPC has been studied at 37.5 degrees C by using unilamellar vesicles of varying sizes. The E/M dip is observable in large unilamellar vesicles; however, the depth of the E/M dip decreases when the vesicle diameter is reduced. When the vesicle diameter is reduced to about 64 nm, the dip becomes shallow and split. This result suggests that the curvature-induced increase in the separation of lipids in the outer monolayer decreases the tendency of regular distribution for pyrene-labeled acyl chains. Regular distribution is believed to arise from the long-range repulsive interaction between Pyr-PC molecules due to the elastic deformation of the lipid matrix around the bulky pyrene moiety. When the radius of curvature becomes small, outer monolayer lipids are more separated. Therefore, pyrene-containing acyl chains fit better into the membrane matrix, which alleviates the deformation of the lattice and diminishes the long-range repulsive interactions between pyrene-containing acyl chains. Furthermore, we have shown a striking difference in the pressure dependence of E/M at critical Pyr-PC mole fractions and at noncritical mole fractions. In the pressure range between 0.001 and 0.7 kbar at 30 degrees C, E/M decreases steadily with increasing pressure at noncritical mole fractions; in contrast, E/M changes little with pressure at critical mole fractions (e.g., 33.3 and 50.0 mol% Pyr-PC). The pressure data suggest that membrane free volume in the liquid crystalline state of the bilayer is less abundant at critical Pyr-PC mole fractions than at noncritical mole fractions.

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High electric field impulses (1-20 kV/cm, 1-20 microseconds) may trigger fusion between adhering cells or lipid vesicles (electrofusion). In this paper a qualitative model of electrofusion is proposed consistent with both electron and light microscopic data. Electrofusion is considered as a multistep process comprising tight membrane-contact formation, membrane electroporation as well as an alternating series of subsequent fast collective and slow diffusive fusion stages. The following sequence of steps is suggested: The electric field pulse enforces (via polarization) a tight contact between the membranes of the cells or vesicles to be fused. During tight-contact formation between the opposing membrane surfaces the membrane-adherent water layers are partially squeezed out from the intermembraneous space. Pores are formed in the double membrane contact area (electroporation) involving lateral diffusion and rotation of the lipid molecules in both adhering membrane parts. With increasing pore density, pore-pore interactions lead to short-range coalescence of double membrane pores resulting in ramified cracks; especially small tongues and loops are formed. At supercritical pore density long-range coalescence of the pores occurs (percolation) producing one large double membrane loop (or tongue) and subsequently one large hole in the contact area. After switching off the electric field, the smaller pores, tongues and loops reseal and water flows back into the intermembraneous space of the double membrane in the contact area. As a consequence of the increasing membrane-membrane separation due to water backflow, cooperative rounding of the edges of remaining larger tongues and holes occurs. This results in the formation of an intercellular cytoplasm bridge (channel) concomitant with the disappearance of the contact line between the fusing cells. The membrane parts surrounded by continuous loop-like cracks may separate from the system and may finally form vesicles. Our electrofusion model comprises a strong linkage between the membrane pore formation by high electric fields (electroporation) and the process of electrofusion. Additionally, both pore-pore interactions as well as protein-protein interactions in the contact area of the fusing cells are explicitly introduced. The model provides a qualitative molecular description of basic experimental observations such as the production of membrane fragments, of smaller inside-out vesicles and the formation of larger intercellular cytoplasm bridges.

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The lateral distribution of N-[10(1-pyrenyl)decanoyl]-sphingomyelin (PyrSPM) and N-[10(1-pyrenyl)decanoyl]-glucocerebroside (PyrGlcCer) was studied in multilamellar vesicles of 1,2-dipalmitoyl-, 1,2-dimyristoyl-, and 1-palmitoyl-2-oleoyl-phosphatidylcholine (DPPC, DMPC, and POPC, respectively) under anaerobic conditions by determining the excimer-to-monomer fluorescence intensity ratio (E/M) as a function of temperature. The E/M(T) curves for PyrSPM and PyrGlcCer in the three phosphatidylcholine matrices are qualitatively similar to the curves reported for 1-palmitoyl-2-[10-(1-pyrenyl)decanoyl]-phosphatidylcholine (PyrPC) in the same three matrix phospholipids (Hresko, R. C., I. P. Sugár, Y. Barenholz, and T. E. Thompson, 1986, Biochemistry, 25:3813-3823). However, there is independent evidence to suggest that sphingomyelin and glucocerebroside are organized in POPC, DPPC, and DMPC in a more complex manner than is PyrPC. In an effort to examine further the relationship between the lateral distribution of the labeled lipid and the shape of an E/M(T) curve, E/M vs. temperature simulations were carried out together with an analysis of the equation that relates E/M to the system parameters. The results indicate that information about the lateral distribution of the pyrene-labeled lipid can be obtained from an E/M(T) curve only for those systems in which the gel to liquid crystalline phase transition temperature of the matrix lipid is higher than that of the pyrene-labeled lipid. However, very little can be known about the system from an E/M(T) curve if the matrix lipid has the lower phase transition temperature.

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The lateral distribution of 1-palmitoyl-2-[10-(1-pyrenyl)decanoyl]phosphatidylcholine (PyrPC) was studied in small unilamellar vesicles of 1,2-dipalmitoyl-, 1,2-dimyristoyl-, and 1-palmitoyl-2-oleoyl-phosphatidylcholine (DPPC, DMPC, and POPC, respectively) under anaerobic conditions. The DPPC and DMPC experiments were carried out over temperature ranges above and below the matrix phospholipid phase transition temperature (Tm). The excimer to monomer fluorescence intensity ratio (E/M) was determined as a function of temperature for the three PyrPC/lipid mixtures. Phase and modulation data were used to determine the temperature dependence of pyrene fluorescence rate parameters in gel and in liquid-crystalline bilayers. These parameters were then used to provide information about excited-state fluorescence in phospholipid bilayers, calculate the concentration of the probe within liquid-crystalline and gel domains in the phase transition region of PyrPC in DPPC, and simulate E/M vs. temperature curves for three systems whose phase diagrams are different. From the simulated curves we could determine the relationship between the shape of the three simulated E/M vs. temperature curves and the lateral distribution of the probe. This information was then used to interpret the three experimentally derived E/M vs. temperature curves. Our results indicate that PyrPC is randomly distributed in pure gel and fluid phosphatidylcholine bilayers.(ABSTRACT TRUNCATED AT 250 WORDS)

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