The long-term goal of this line of investigation is to develop efficient protein transfer technologies for engineering cell surfaces; experimental and therapeutic applications are envisioned. To this end, this proposal investigates the potential application of glycosyl-plasmanylinositol (GPI) anchors to protein transfer. The central premise is that natural, as well as artificial, GPI-modified polypeptides can be efficiently reincorporated into cellular membranes, and once reincorporated, retain critical functions of their respective parental proteins. Moreover, an additional premise is that the artificial linking of GPI anchors to polypeptides can confer to these polypeptides novel functional capabilities. The preliminary data exemplify the way in which chimeric gene transfer can be applied for artificial GPI modification and for conveying selected functional properties to target polypeptides. The next logical step is to move from gene to protein transfer as the means for delivering GPI-modified proteins to the cell surface. The sequential series of specific aims of this proposal are a combination of technological and biological objectives.
Specific Aim #1 is directed toward the quantitative production of GPI- modified polypeptides using genetic over-expression systems.
Specific Aim #2 seeks to devise efficient methodologies for membrane reincorporation of GPI-modified polypeptides.
Specific Aims #3 and #4 address biological features of GPI-modified polypeptides that have been introduced into membranes by protein transfer. The proposed investigations will advance GPI anchor studies by facilitating the use of protein transfer, as a complementary technology to gene transfer, for introducing GPI-anchored proteins into cell surfaces. Insights into the functional repertoire of GPI-anchored proteins may ensue. Additionally, these studies may lead to new approaches for engineering the cell surface phenotype of cells to be used in cell transfer therapeutics.
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