Membrane arrays and methods of manufacture
The invention relates to G protein-coupled receptor (GPCR) microarrays on porous substrates for structural or functional analyses of GPCRs, and methods of preparing porous substrate surfaces for receiving membranes that comprise GPCRs. In one embodiment, a GPCR microarray of the invention comprises a membrane adhered to an upper surface of a porous substrate, the membrane spanning across a plurality of pores on the porous substrate to form a plurality of cavities having sufficient geometry to permit entry of assay reagents into each cavity, thereby allowing access of assay reagents to both sides of GPCR in the membrane.
1. A method for identifying a modulator of a membrane protein, comprising:
contacting a membrane with a candidate molecule, said membrane comprising said membrane protein immobilized on a porous substrate comprising a non-porous base region and a porous region adjacent to the non-porous base region, and both sides of said membrane being accessible to an assay agent; and
detecting a biological function of said membrane protein, wherein a change in biological function in the presence of said candidate molecule as compared to that in the absence of said candidate molecule is indicative of the capability of said candidate molecule to modulate said membrane protein.
2. The method of claim 1 , wherein said membrane protein is selected from the group consisting of a G protein coupled receptor, an ion channel, a kinase receptor, and a transporter.
3. The method of claim 1 , wherein said membrane is a cellular membrane.
4. The method of claim 1 , wherein said membrane protein is a G protein coupled receptor, and said membrane comprises a G protein or a subunit thereof, and wherein said function is detected by measuring activation of said G protein or subunit by said G protein coupled receptor.
5. The method of claim 4 , comprising contacting said membrane with (1) a ligand of said G protein coupled receptor and (2) a GTP analogue capable of binding to said G protein or subunit upon activation of said G protein or subunit.
6. The method of claim 5 , wherein said candidate molecule is an agonist of said G protein coupled receptor.
7. The method of claim 5 , wherein said candidate molecule is an antagonist of said G protein coupled receptor.