Differential effects of surfactant protein A on regional organization of phospholipid monolayers containing surfactant protein B or C

Epifluorescence microscopy combined with a surface balance was used to stud y monolayers of dipalmitoylphosphatidylcholine (DPPC)/egg phosphatidylglyce rol (PG) (8.2, mol/mol) plus 17 wt % SP-B or SP-C spread on subphases conta ining SP-A in the presence or absence of 5 mM Ca2+. Independently of the pr esence of Ca2+ in the subphase, SP-A at a bulk concentration of 0.68 mu g/m l adsorbed into the spread monolayers and caused an increase in the molecul ar areas in the films. Films of DPPC/PG formed on SP-A solutions showed a p ressure-dependent coexistence of liquid-condensed (LC) and liquid-expanded (LE) phases. Apart from these surface phases, a probe-excluding phase, like ly enriched in SP-A, was seen in the films between 7 mN/m less than or equa l to pi less than or equal to 20 mN/m. In monolayers of SP-B/(DPPC/PG) spre ad on SP-A, regardless of the presence of calcium ions, large clusters of a probe-excluding phase, different from probe-excluding lipid LC phase, appe ared and segregated from the LE phase at near-zero surface pressures and co existed with the conventional LE and LC phases up to similar to 35 mN/m. Va rying the levels of either SP-A or SP-B in films of SP-B/SP-A/(DPPC/PG) rev ealed that the formation of the probe-excluding clusters distinctive for th e quaternary films was influenced by the two proteins. Concanavalin A in th e subphase could not replace SP-A in its ability to modulate the textures o f films of SP-B/(DPPC/PG), In films of SP-C/SP-A/(DPPC/PG), in the absence of calcium, regions consisting of a probe-excluding phase, likely enriched in SP-A, were detected at surface pressures between 2 mN/m and 20 mN/m in a ddition to the lipid LE and LC phases. Ca2+ in the subphase appeared to dis perse this phase into tiny probe-excluding particles, likely comprising Ca2 +-aggregated SP-A. Despite their strikingly different morphologies, the fil ms of DPPC/PG that contained combinations of SP-B/SP-A or SP-C/SP-A display ed similar distributions of LC and LE phases with LC regions occupying a ma ximum of 20% of the total monolayer area. Combining SP-A and SP-B reorganiz ed the morphology of monolayers composed of DPPC and PG in a Ca2+-independe nt manner that led to the formation of a separate potentially protein-rich phase in the films.