Characterization of a signaling molecule assembly region within the cell membrane

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Characterization of a signaling molecule assembly region(raft) within the cell membrane in the brain tissue resulted in the following findings :1. Major components were identified as, tubulin, actin, NAP-22, GAP-43, and several trimeric G proteins, using western blotting and partial amoni acid sequencing analysis.2. Concentration of several GPI-anchoed proteins, such as F3/contactin, T-cadherin, OBCAM, and neurotrimin was found. In addition, a novel GPI-anchored protein was idenfied and cloned. This protein, named kilon, showed a wide distribution in the brain cortex.3. Production of monoclonal antobodies using raft as an immunogen resulted in the identification of a protein focalized in raft. This protein was identified as paralemin from its amino acid sequence.4. Lipid analysis of raft showed the enrichment of phosphatidylcholine and cholesterol in this fraction. Extraction of cholesterol using methy-cyclodextrin caused specific solubilization of NAP-22. Further studies showed the binding of NAP-22 to a cholestrol containing liposome.