| Abstract
| - Positioned at the C-terminus of many eukaryotic proteins, the glycosylphosphatidylinositol (GPI)anchor is a posttranslational modification that anchors the modified proteins in the outer leaflet of the plasmamembrane. GPI-anchored proteins play vital roles in signal transduction, the vertebrate immune response,and the pathobiology of trypanosomal parasites. While many GPI-anchored proteins have been characterized, the biological functions of the GPI anchor have yet to be elucidated at a molecular level. We synthesizeda series of GPI-protein analogues bearing modified anchor structures that were designed to dissect thecontribution of various glycan components to the GPI-protein's membrane behavior. These anchor analogueswere similar in length to native GPI anchors and included mimics of the native structure's three domains.A combination of expressed protein ligation and native chemical ligation was used to attach these analoguesto the green fluorescent protein (GFP). These modified GFPs were incorporated in supported lipid bilayers,and their mobilities were analyzed using fluorescence correlation spectroscopy. The data from theseexperiments suggest that the GPI anchor is more than a simple membrane-anchoring device; it also mayprevent transient interactions between the attached protein and the underlying lipid bilayer, thereby permittingrapid diffusion in the bilayer. The ability to generate chemically defined analogues of GPI-anchored proteinsis an important step toward elucidating the molecular functions of this interesting post-translationalmodification.
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