Intermolecular interaction of phosphatidylinositol with the lipid raft molecules sphingomyelin and cholesterol

Abstract

Diacylphosphatidylinositol (PI) is the starting reactant in the process of phosphatidylinositide-related signal transduction mediated through the lipid raft domain. We investigated intermolecular interactions of PI with major raft components, sphingomyelin (SM) and cholesterol (Chol), using surface pressure-molecular area (π-A) isotherm measurements. The classical mean molecular area versus composition plot showed that the measured mean molecular areas are smaller in PI/Chol mixed monolayers and larger in PI/SM mixed monolayers than those calculated on the basis of the ideal additivity. These results indicate that PI interacts attractively with Chol and repulsively with SM. In addition, we energetically evaluated the interaction of PI with SM/Chol mixtures and found that the mixing energy of PI/SM/Chol ternary monolayers decreased as the molar ratio of Chol to SM increased. In order to quantitatively analyze the distribution of PI we calculated the chemical potentials of mixing of PI into the SM/Chol mixed monolayer and into the dioleoylphosphatidylcholine (DOPC) monolayer, which was used as a model for the fluid matrix, on the basis of partial molecular area analysis. Analysis using the chemical potential of mixing of PI suggested that partition of PI molecules between these two monolayers can be changed by a factor of about 1.7 in response to change in Chol molar fraction in the SM/Chol mixed monolayer from 0.3 to 0.6 when the concentration of PI in the DOPC monolayer is kept constant at 7 mol%.

Keywords: Gibbs mixing energy; chemical potential of mixing; model monolayer; molecular distribution; π–A isotherm.

Figure 1.

Intermolecular interaction in the PI/SM monolayer system. (a) π–A isotherms of pure PI, pure SM and PI/SM mixed monolayers on the water subphase at 25±0.1°C. The molar fractions of PI, X_PI, are indicated in the figure; 0 (SM), 0.3, 0.5, 0.7, 0.9 and 1.0 (PI). The isotherms with X_PI= 0.5 and 0.9 were nearly superposed upon those of X_PI= 0.7 and 1.0, respectively. (b) Mean molecular area versus composition analysis at 30 mN/m. The dotted line represents area additivity for ideal mixing of two components (Eq. (1)). (c) Areal compressional modulus ( $C_{\text{s}}^{-1}$) versus composition analysis. The $C_{\text{s}}^{-1}$ values at 30 mN/m were calculated from equation (4). The solid line represents ideal additivity of compressibility (see Materials and methods and Eq. (5)).

Figure 2.

Intermolecular interaction in the PI/Chol monolayer system. (a) π–A isotherms of pure PI, pure Chol and PI/Chol mixed monolayers on the water subphase at 25±0.1°C. The molar fractions of PI, X_PI, are indicated in the figure; 0 (Chol), 0.3, 0.5, 0.7, 0.9 and 1.0 (PI). (b) Mean molecular area versus composition analysis at 30 mN/m. The dotted line represents area additivity (Eq. (1)). (c) Areal compressional modulus ( $C_{\text{s}}^{-1}$) versus composition analysis. The $C_{\text{s}}^{-1}$ values at 30 mN/m were calculated from equation (4). The solid line represents ideal additivity of compressibility (see Materials and methods and Eq. (5)).

Figure 3.

Intermolecular interaction in the PI/DOPC monolayer system. (a) π–A isotherms of pure PI, pure DOPC and PI/DOPC mixed monolayers on the water subphase at 25±0.1°C. The molar fractions of PI, X_PI, are indicated in the figure; 0 (DOPC), 0.3, 0.5, 0.7, 0.9 and 1.0 (PI). (b) Mean molecular area versus composition analysis at 30 mN/m. The dotted line represents area additivity (Eq. (1)). The molecular area of DOPC (0.64 nm²) is larger than that of PI (0.55 nm²) at 30 mN/m.

Figure 4.

Excess mixing energy of PI and SM/Chol mixtures (SCm), ΔG_ex^SCm, as a function of X_PI. The value of ΔG_ex^SCm was calculated by integration of area deviation (ΔA) over the surface pressure (Eq. (7)). The molar ratios of Chol in the SCm, r_Chol, are 0 (filled inverted triangle), 0.3 (filled square), 0.6 (filled circle), 0.9 (filled diamond) and 1.0 (filled triangle).

Figure 5.

Analysis of partial molecular area of PI at given X_PI, A_PI(X_PI). Mean molecular area versus X_PI plot for the PI/SCm (r_Chol= 0.9) mixed monolayer at 30 mN/m is shown as an example for calculation of A_PI(X_PI). The mean molecular area data were fitted to a quadratic function, f(X_PI) (solid line). The value of A_PI(X_PI) obtained as the intercept at X_PI= 1.0 of the tangent to the fitted curve f(X_PI); the calculation in the case of X_PI= 0.3 (arrow) is shown in the figure. ΔA_PI= A_PI(X_PI)–A_PI(1), where A_PI(1) is the molecular area of pure PI.

Figure 6.

Chemical potential of mixing of PI into the SCm with a fixed r_Chol-value (Δμ_PI^SCm) and into the DOPC monolayer (Δμ_PI^DOPC) as a function of X_PI. As shown in equation (10), Δμ_PI consists of the ideal term and the integration of ΔA_PI over the surface pressure (π=0–30 mN/m). The molar ratios of Chol in the SCm, r_Chol, are 0 (filled inverted triangle), 0.3 (filled square), 0.6 (filled circle), 0.9 (filled diamond) and 1.0 (filled triangle). The open stars correspond to Δμ_PI^DOPC. Note that Δμ_PI^SCm (r_Chol= 0.6) at X_PI= 0.12 (arrow b) equals to Δμ_PI^DOPC at X_PI= 0.07 (arrow a). See text for details.

Figure 7.

Chol-induced redistribution of PI molecules between the SCm and the fluid matrix, speculated from the results obtained in the model monolayer systems. When r_Chol∼0.3, PI may be equally distributed between the fluid matrix and the SCm (upper). The PI molecules will be gradually transferred from the fluid matrix to the SCm as r_Chol increases (lower).