Effect of Hydrophobic Surfactant Proteins SP-B and SP-C on Binary Phospholipid Monolayers

Abstract

In situ external reflection infrared spectroscopy at the air-water interface was used to study the influence on phospholipid structure of an endogenous mixture of the two hydrophobic surfactant proteins, SP-B and SP-C, which are thought to play pivotal roles in the adsorption and function of pulmonary surfactant. Mixtures studied were 1:1, 2:1, and 7:1 (mol:mol) DPPC-d(62):DPPG, and 7:1 DPPC-d(62):DOPG, alone and in the presence of 0.5-10 wt % mixed SP-B/C purified chromatographically from calf lung surfactant extract. Perdeuteration of DPPC produced a shift in vibrational frequencies so that it could be differentiated spectroscopically from the phosphoglycerol component in the surface monolayer. CH(2) antisymmetric and symmetric stretching bands ( approximately 2920 and 2852 cm(-1)) along with the analogous CD(2) stretching bands ( approximately 2194 and 2089 cm(-1)) were analyzed, and band heights and peak wavenumber positions were assessed as a function of monolayer surface pressure. Small, near-physiological contents of 1-2 wt % SP-B/C typically produced the maximum observed spectroscopic effects, which were abolished at high protein contents of 10 wt %. Analysis of CH(2) and CD(2) stretching bands and C-H/C-D band height ratios indicated that SP-B/C affected PC and PG lipids differently within the surface monolayer. SP-B/C had preferential interactions with DPPG in 1:1, 2:1, and 7:1 DPPC-d(62):DPPG films that increased its acyl chain order. SP-B/C also interacted specifically with DOPG in 7:1 DPPC-d(62):DOPG monolayers, but in this case an increase in CH(2) band heights and peak wavenumber positions indicated a further disordering of the already fluid DOPG acyl chains. CD(2) band height and peak wavenumber analysis indicated that SP-B/C had no significant effect on the structure of DPPC-d(62) chains in 7:1 films with DPPG or DOPG, and had only a slight tendency to increase the acyl chain order in 1:1 films of DPPC-d(62):DPPG. SP-B/C had no significant effect on DPPC-d(62) structure in films with DOPG. Infrared results also indicated that interactions involving SP-B/C and lipids led to exclusion of PC and PG lipids from the compressed interfacial monolayer, in agreement with our previous report on the phase morphology of lipid monolayers containing 1 wt % SP-B/C.

Protein–lipid interactions and surface activity in the pulmonary surfactant system

“…4). The disposition of SP-B in lipid bilayers and monolayers as well as the effect produced on the lipids has been also analyzed by many different techniques such as 2 H NMR (Morrow et al, 1993(Morrow et al, , 2004, spin resonance (Perez-Gil et al, 1995), infrared spectroscopy (Brockman et al, 2003), atomic force microscopy (Cruz et al, 2004) or epifluorescence microscopy (Cruz et al, 2000).…”

Distinct effects of SP-B and SP-C on the uptake of surfactant-like liposomes by alveolar cells in vivo and in vitro

“…The increment in uptake caused by SP-B at the high proteinto-lipid ratio could also be explained by changes in lipid conformation of the liposomes because SP-B is able to change lipid conformation of surfactant (5).…”

High‐resolution Studies of Lung Surfactant Collapse¶

“…The first plateau observed is taken as evidence of removal of material from the monolayer. Previous studies propose that the material removed may be squeezed out into the subphase or form surface‐associated aggregates ( 18–21 ). Similar nonhorizontal plateaus are observed in many systems, and current theoretical investigations treat this behavior as arising from surface micelle formation ( 22,23 ).…”

“…SP‐C also exhibits the same partitioning. It has also been shown that SP‐B and SP‐C preferentially associate with anionic lipids ( 18,19 ). It then follows that the bright, expanded regions seen in the fluorescent images of the LS are also rich in anionic lipids.…”