Development of Highly Sensitive Bilayr Lipid Membrane Sensors Based on Membrane Proteins

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Fundamental studies aiming at generalization of bilayr lipid membrane sensors, which exploit remarkable signal amplification function of ion channel proteins for highly sensitive sensing of target compounds, were carried out and the following results were obtained.1.Glutamate receptor ion channel protein (GluR), purified from postsynaptic membranes of rat brain, was incorporated into a planar bilayr lipid membrane, and its agonist selectivity was evaluated for several representative agonists under the conditions in which only the NMDA subtype was active. The agonist selectivity based on the total ion permeated through the channels (Ca^<2+> and Na^+) was lower than that of the binding assay, although the order of the selectivity was the same for the two systems. In addition, the agonist selectivity based on the permeation of Ca^<2+> ions was evaluated by thin layr potentiometry using liposomes incorporated with the GluR proteins. A selectivity similar to that for planar bilayr lipid mem … More branes was observed for the NMDA subtype. The present approach based on the transmembrane signals from the GluR proteins (total ions permeated through the membrane) affords a new method for evaluating a biocompatible selectivity, which is different from the binding affinity used so far to evaluate the agonist selectivities of channel proteins.2.Fabrication of a hybrid-type bilayr lipid membrane, incorporated with voltage-sensitive sodium channel proteins (from electric organ of electric eel) with artificial receptor molecules capable of generating guest-induced membrane potential changes, was attempted. Planar bilayr lipid membranes incorporated with the above ion channel protein and valinomycin were prepared. Although the incorporated channel proteins kept ion channel activity, addition of K^+ ion in low concentrations caused a marked decrease in the membrane resistance, indicating the necessity of reinvestigation of the experimental system to make it able to measure the changes in channel currents involving membrane potential changes. Less