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SP-A Permeabilizes Lipopolysaccharide Membranes by Forming Protein Aggregates that Extract Lipids from the Membrane

Olga Cañadas ^* , Ignacio García-Verdugo ^*, Kevin M. W. Keough  and Cristina Casals ^* 

^* Department of Biochemistry and Molecular Biology I, and Centro de Investigaciones Biomédicas en Red Enfermedades Respiratorias, Complutense University of Madrid, Madrid, Spain; and Department of Biochemistry and Discipline of Pediatrics, Memorial University of Newfoundland, St. John's, Newfoundland, Canada

Correspondence: Address reprint requests to Cristina Casals, PhD, Dept. of Biochemistry and Molecular Biology I, Faculty of Biology, Complutense University of Madrid, 28040 Madrid, Spain. Tel.: 34-91-3944261; Fax: 34-91-3944672; E-mail: ccasalsc{at}bio.ucm.es.

Surfactant protein A (SP-A) is known to cause bacterial permeabilization. The aim of this work was to gain insight into the mechanism by which SP-A induces permeabilization of rough lipopolysaccharide (Re-LPS) membranes. In the presence of calcium, large interconnected aggregates of fluorescently labeled TR-SP-A were observed on the surface of Re-LPS films by epifluorescence microscopy. Using Re-LPS monolayer relaxation experiments at constant surface pressure, we demonstrated that SP-A induced Re-LPS molecular loss by promoting the formation of three-dimensional lipid-protein aggregates in Re-LPS membranes. This resulted in decreased van der Waals interactions between Re-LPS acyl chains, as determined by differential scanning calorimetry, which rendered the membrane leaky. We also showed that the coexistence of gel and fluid lipid phases within the Re-LPS membrane conferred susceptibility to SP-A-mediated permeabilization. Taken together, our results seem to indicate that the calcium-dependent permeabilization of Re-LPS membranes by SP-A is related to the extraction of LPS molecules from the membrane due to the formation of calcium-mediated protein aggregates that contain LPS.