Anionic Phospholipids change the Effect of the Hydrophobic Surfactant Proteins on Structures of Hexagonal Lipids

Available evidence suggests that the hydrophobic surfactant proteins (SPs), SP-B and SP-C, accelerate adsorption of surfactant vesicles to an air/water interface by promoting formation of a negatively curved rate-limiting structure. In support of this model, the proteins induce several phosphatidylethanolamines to form inverse bicontinuous cubic phases, in which each leaflet has negative saddle-like curvature analogous to the hypothetical intermediate. The proteins could promote formation of cubic phases by changing spontaneous curvature (c0) of lipids, which would be reflected in the dimensions of the inverse hexagonal (HII) phase. With 1,2-dioleoyl phosphatidylethanolamine (DOPE), the SPs had no effect on the size of the HII phase, suggesting a constant c0. The study, however, lacked anionic phospholipids, which constitute ∼10% (mol:mol) of phospholipids in lung surfactant, and could engage in selective interactions with the cationic SPs. In this work, we used small-angle X-ray scattering to examine how SPs affect structures formed by DOPE mixed with 10% (mol:mol) anionic 1,2-dioleoyl phosphatidylglycerol (DOPG). With DOPG, the HII lattice-constant (a0) decreased in a dose-dependent manner with increasing levels of protein. This change could be caused by specific interactions between the SPs and DOPG, or nonspecific interactions between cationic SPs and the anionic phosphate group of DOPG. To determine whether the observed change in a0 was caused by nonspecific electrostatic effects, the measurements were repeated on samples prepared in buffered electrolyte. In the presence of counterions, the effect of SPs on a0 was significantly diminished. These results suggest that the change in the a0 for DOPE:DOPG in the absence of counterions is caused by nonspecific electrostatic interactions between the positively-charged proteins and anionic phospholipids, and are unlikely to play a major role in physiological media.Acknowledgements: Stanford Synchrotron Radiation Lightsource; NIH.