Lipids play an important structural and metabolic role in all living systems. Consequently the effective shuttling of these water insoluble molecules within the aqueous environment of an organism is of central biological importance. The overall long term objective of this research to understand on a molecular level the interactions between lipids and the proteins specifically designed for their transport. This is particularly relevant since impairment of such transport in humans can lead to serious cardio- vascular problems. For the structural studies proposed here, two insect lipid transport proteins have been chosen: apolipophorin-III isolated from Locusta migratoria and insecticyanin isolated from Manduca sexta L. These proteins were selected for study because they are believed to have different three-dimensional structural motifs for binding lipids and because they have been well studied using both biochemical and biophysical techniques. Initially the high resolution structures will be determined by x-ray crystallographic methods. Subsequently, by using site-directed mutagenesis techniques, """"""""mutants"""""""" with modified lipid binding capacities will be produced and their three-dimensional structures determined. In the case of insecticyanin, its molecular structure to 2.6 A resolution has been solved in this laboratory and the study will be extended to 2.2 A resolution. Especially significant to this proposal is the suggestion that apolipoprotein D, which comprises 5% of the human high-density lipoprotein, may have a similar molecular fold as observed in insecticyanin. With respect to apolipophorin-III, the proposed research is particularly important since it is the first apolipophorin from either vertebrate or invertebrate sources to be crystallized in a form suitable for a high resolution x-ray analysis. Consequently, while the proposed research focuses on insect systems, the work will also provide valuable insight into mammalian lipid transport pathways as well.
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