LIPID CONTROL OF MEMBRANE PROTEIN ORGANIZATION

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DESCRIPTION (Adapted from abstract): The major focus of this laboratory to determine how the structures of lipids and proteins are altered upon their mutual interaction in a series of model and native membranes of increasing complexity, and how these structural modifications are related to function. The investigators have developed FT-IR experiments that yield quantitative information about membrane lipid conformational and orientational order and applied this technology to a physiologically relevant experimental paradigm, i.e., monolayers in situ at the air/water (A/W) interface. This experiment, a model for the pulmonary surfactant system, may provide an understanding as to how the surfactant components are organized at the air/alveolar lining. This information in turn may permit the rational design of. therapeutic agents for diseases such as respiratory distress syndrome. Two specific aims, each with a physical and biological/biomedical component, will be pursued during the coming funding period: (Aim 1, physical): IR reflection-absorption spectroscopy (IRRAS) will be developed for efficient measurement of conformations, head group interactions and tilt angles of phospholipid acyl chains and secondary structures and tilt angles of ordered regions (a-helix, Beta-sheet) of peptides in aqueous monolayers in situ at the A/W interface. The investigators will also continue to develop mathematical methods for reflected intensities for quantitative interpretation of results. (Aim 1. biological): The investigators will explore the molecular level ramifications of the "squeeze-out" hypothesis widely assumed to be the mechanism of lung function. To achieve this, they will measure the conformations and orientations of specific segments within the pulmonary surfactant proteins SP-B and SP-C in monolayer and bulk phase environments. Finally they will utilize the combination of IRRAS and Brewster Angle Microscopy to monitor interactions of SP-A, -B, and -C with lipids at the molecular level (IRRAS) and the 10 u-M scale (BAM) in monolayers. (Aim 2, physical) The investigators will apply the novel techniques of IR microscopic imaging to acquire spectrally and spatially resolved IR images from heterogeneous systems (e.g., ceramide/fatty acids/cholesterol and other phase separated lipid mixtures) as the first detailed demonstration of the utility of this technology. (Aim 2, biological) The investigators will extend IR imaging to physiological systems using stratum corneum as a model since the currently accepted molecular description of the skin barrier function postulates the existence of large domains of highly ordered lipidic phases. They will directly evaluate the applicability of domain mosaic model that is the currently accepted model of the stratum corneum.