title: Effect of hydrophobic surfactant protein SP-C on binary phospholipid monolayers. Molecular machinery at the air/water interface.
abstract: Fluorescent and modified dark-field microscopies were used to investigate the phase behavior of physiologically relevant lipid/protein monomolecular films containing surfactant protein C(SP-C). Synthetic human SP-C(1-34) was labeled at its N-terminus using the fluorescent probe 6-(((4(4,4-difluoro-5-(2-thienyl)-4-bora-3a,4a-diaza-s-indacene-3-yl)phenoxy)acetyl)amino)hexanoic acid (BODIPY/TR-X). Using dual fluorescent labeling (lipid and protein) in the monolayers, we have correlated (at physiologically small concentrations of the protein) the lipid phase separation and protein distribution in situ. A comparison of the lipid and protein dye fluorescent micrographs indicates that SP-C(1-34) is preferentially associated with the disordered lipid phase. Three concepts arise from our results. (1) The presence of SP-C alone does not result in the complete dissolution of condensed phase domains in a fashion that we have previously reported for the entire hydrophobic surfactant protein (SP-B/C) fraction (Biophys. J. 77 (1999) 903). Rather, the use of relatively high amounts ( approximately 10 wt.%) of the labeled SP-C protein is needed to reproduce the fluorescence monolayer morphology previously observed for small concentrations ( approximately 1 wt.%) of the natural SP-B/C mixture. (2) Scattered light, dark-field microscopy performed using grazing angle laser illumination reveals the presence of surface-associated, three-dimensional (3D) structures of micrometer-sized dimensions when the labeled BODIPY/TR-X:SP-C(1-34) protein is included in the monolayer, as previously observed with the naturally isolated SP-B/C mixture. The 3D structures are associated exclusively with the presence of the SP-C protein in disordered monolayer phases. (3) To explain these results, we have derived a molecular model accounting for the structure and physico-chemical properties of the SP-C protein in terms of its energetics. The molecular events involved in the SP-C-mediated production of the 3D surface particles are explained using the analogy of a simple molecular machine, namely a loaded spring. This interpretation is supported by an energetic analysis that suggests the major factor contributing to the formation of the 3D particles is the energy liberated by re-expansion of the surrounding phospholipid film into the area vacated by the SP-C protein as it re-orients away from the surface.
authors.affiliation: Department of Chemistry, University of Georgia, Athens, GA 30602-2556, USA.
authors.forename: Peter
authors.initials: P
authors.lastname: Krüger
authors.forename: John E
authors.initials: JE
authors.lastname: Baatz
authors.forename: Richard A
authors.initials: RA
authors.lastname: Dluhy
authors.forename: Mathias
authors.initials: M
authors.lastname: Lösche
mesh_terms: D000388:Air; D000465:Algorithms; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D006801:Humans; D008027:Light; D008567:Membranes, Artificial; D008856:Microscopy, Fluorescence; D010743:Phospholipids / Q000737:chemistry*; D037721:Pulmonary Surfactant-Associated Protein C / Q000737:chemistry*; D012542:Scattering, Radiation; D013055:Spectrophotometry, Infrared; D013499:Surface Properties
chemical_list: D008567:Membranes, Artificial; D010743:Phospholipids; D037721:Pulmonary Surfactant-Associated Protein C