COMBINATIONS OF FLUORESCENTLY LABELED PULMONARY SURFACTANT PROTEINS SP-B AND SP-C IN PHOSPHOLIPID FILMS

Hydrophobic pulmonary surfactant (PS) proteins B (SP-B) and C (SP-C) m odulate the surface properties of PS lipids. Epifluorescence microscop y was performed on solvent-spread monolayers of fluorescently labeled porcine SP-B (R-SP-B, labeled with Texas Red) and SP-C (F-SP-C, labele d with fluorescein) in dipalmitoylphosphatidylcholine (DPRC) (at prote in concentrations of 10 and 20 wt%, and 10 wt% of both) under conditio ns of cyclic compression and expansion. Matrix-assisted laser desorpti on/ionization (MALDI) spectroscopy of R-SP-B and F-SP-C indicated that the proteins were intact and labeled with the appropriate fluorescent probe. The monolayers were compressed and expanded for four cycles at an initial rate of 0.64 Angstrom(2) . mol(-1) . s(-1) (333 mm(2) . s . (-1)) up to a surface pressure pi approximate to 65 mN/m, and pi-are a per residue (pi-A) isotherms at 22 +/- 1 degrees C were obtained. Th e monolayers were microscopically observed for the fluorescence emissi on of the individual proteins present in the film lipid matrix, and th eir visual features were video recorded for image analysis. The pi-A i sotherms of the DPPC/protein monolayers showed characteristic ''squeez e out'' effects at pi approximate to 43 mM/m for R-SP-B and 55 mN/m fo r F-SP-C, as had previously been observed for monolayers of the native proteins in DPPC. Both proteins associated with the expanded (fluid) phase of DPPC monolayers remained in or associated with the monolayers at high pi (-65 mN/m) and redispersed in the monolayer upon its reexp ansion. At comparable pi and area/molecule of the lipid, the proteins reduced the amounts of condensed (gel-like) phase of DPPC monolayers, with F-SP-C having a greater effect on a weight basis than did R-SP-B. In any one of the lipid/protein monolayers the amounts of the DPPC in condensed phase were the same at equivalent pi during compression and expansion and from cycle to cycle. This indicated that only minor los s of components from these systems occurred between compression-expans ion cycles. This study indicates that hydrophobic PS proteins associat e with the fluid phase of DPPC in films, some proteins remain at high surface pressures in the films, and such lipid-protein films can still attain high pi during compression.