INTERACTIONS OF HYDROPHOBIC LUNG SURFACTANT PROTEINS SP-B AND SP-C WITH DIPALMITOYLPHOSPHATIDYLCHOLINE AND DIPALMITOYLPHOSPHATIDYLGLYCEROL BILAYERS STUDIED BY ELECTRON-SPIN-RESONANCE SPECTROSCOPY

Hydrophobic surfactant-associated proteins SP-B and SP-C have been isolated from porcine lungs and reconstituted in multilamellar vesicles of dipalmitoylphosphatidylcholine (DPPC) or dipalmitoylphosphatidylglycerol (DPPG) containing different phospholipid spin probes, in order to characterize the lipid-protein interactions by electron spin resonance (ESR) spectroscopy. Both proteins caused a significant increase in the outer hyperfine splittings of all the ESR spectra, indicating that SP-B and SP-C reduce the mobility of the phospholipid acyl chains. The more hydrophobic SP-C had greater effects on phospholipid bilayers than did SP-B. The effect was saturated at protein/lipid ratios of 20% and 30% (w/w) for SP-B and SP-C, respectively, in bilayers of DPPC. SP-B and SP-C increased the ordering and decreased the mobility of the lipid acyl chains in both DPPC and DPPG bilayers in the fluid phase, without affecting the gel phase on the conventional ESR time scale. On the other hand, both proteins induced a more homogeneous distribution of the phospholipid spin probes in the gel phase of DPPC. The selectivity of the interaction of SP-B and SP-C with different phospholipid species was determined from the ESR spectra of spin-labeled phospholipids with different headgroups in host bilayers of either DPPC or DPPG. SP-B showed a general preference to interact with negatively charged phospholipids, which was modulated in an ionic strength-dependent manner. At near-physiological ionic strength, SP-B showed selectivity for phosphatidylglycerol. SP-C did not show well-defined selectivity for a particular class of phospholipid, but it did show some preference for phosphatidic acid, which was partially abolished at physiological ionic strength. Finally, studies of the effects of the hydrophobic surfactant proteins on the ionization equilibrium of a stearic acid spin probe in bilayers of DPPC showed that both proteins altered the structure at the bilayer surface, causing an increase in the surface polarity. Such changes could be related to the changes in tensoactive properties induced by these proteins in surfactant phospholipids.