LIPID CONTROL OF MEMBRANE PROTEIN ORGANIZATION

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The long-term objective of this laboratory is to determine how the structures of lipids and proteins are altered upon their mutual interaction in a hierarchy of model membrane systems of increasing complexity, and how these structural modifications are related to function. The developed protocols can be transferred to more complex and/or technically difficult but physiologically more relevant experimental paradigms such as monolayers in situ at the A/W interface. General principles extracted from specific examples strengthen the basis for understanding the organization of biological interfaces and how these are altered during pathological states. The specific aim for the MBRS project has a physical and a biological/biomedical component, which will be pursued during the next funding period; The physical aim is to develop FT-IR technology for the direct molecular characterization of conformational and orientational order of both lipids and proteins in situ in monolayers at the A/W interface. We have constructed a novel external reflection (IRRAS) apparatus that has permitted us to acquire the first IR spectra of protein monolayer films at the A/W interface, and thus obtain secondary structure and orientation information. We will continue to develop and improve both the apparatus and the optical spectroscopic models for quantitative interpretations of results. The biomedical application of this technology will be to evaluate structure/function relationships in a physiologically essential yet biochemically manageable system, lung surfactant. IRRAS provides a unique means to test the major hypothesis formulated to describe the molecular mechanism of surfactant function, the "squeeze-out" hypothesis. This hypothesis requires that the major phospholipid component of the surfactant (DPPC) becomes enriched at the major phospholipid component of the surfactant (DPPC) becomes enriched at the surface during successive expansion-compression cycles in vitro (exhalation-inhalation cycles in vivo), to produce the requisite low surface tension at the air-water interface (air-alveolar interface in vivo). We will address several aspects of the hypothesis, including its occurrence, its dependence on lipid structure and conformation, the roles of the surfactant proteins SP-B and SP-C, the effects of compression rates, modification of the subphase, etc. as they alter squeeze-out parameters.