Assays for screening compounds which interact with cation channel proteins, mutant prokaryotic cation channel proteins, and uses thereof

Abstract

Assays for screeing potential drugs or agents that can interact and potentially bind to cation channel proteins, and potentially have uses in treating conditions related to the function of cation channel proteins is provided, along with prokaryotic cation channel proteins mutated to mimic eukaryotic cation channels, which can then be used in assays of the present invention.

Description

CROSS REFERENCE TO RELATED APPLICATIONS [0001] This application is a continuation in part of copending U.S. application Ser. No. 09 / 054,347 filed on Apr. 2, 1998, which is a continuation in part of copending U.S. application Ser. No. 09 / 045,529 filed on Mar. 20, 1998, wherein both U.S. Ser. Nos. 09 / 054,347 and 09 / 045,529 are hereby incorporated by reference in their entireties.GOVERNMENT RIGHTS CLAUSE [0002] The research leading to the present invention was supported in part with National Institutes of Health Grant GM 43949. The government may have rights in the invention.FIELD OF INVENTION [0003] The present invention relates to a crystal of a cation channel protein, and methods of using such a crystal in screening potential drugs and therapeutic agents for use in treating conditions related to the function of such channels in vivo. BACKGROUND OF INVENTION [0004] Although numerous types of channel proteins are known, the main types of ion channel proteins are characterized by the m...

AI Technical Summary

Benefits of technology

[0014] There is provided, in accordance with the present invention, a method of preparing a functional cation channel protein for use in an assay for screening potential drugs or other agents which interact with a cation channel protein, which permits the screening of potential drugs or agents that may be used as potential therapeutic agents in treating conditions related to the function of cation channel proteins in vivo.

[0019] Moreover, numerous solid phase resins to which a functional cation channel protein can be conjugated have applications in a method of preparing a functional cation channel protein for use in an assay, as described above. For example, a solid phase resin comprising insoluble polystyrene beads, PVDF, polyethylene glycol, or a cobalt resin, to name only a few have application in the present invention. Preferably, a cation channel protein is conjugated to a cobalt resin at a protein to resin ratio that allows for saturation of the resin with the cation channel protein. Moreover, after conjugation, the cobalt resin is preferably used to line a column having a volume of about 1 ml.

[0021] Subsequently, the cation channel protein is removed from the solid phase resin, and analyzed to determine whether the potential drug or agent is bound thereto. Numerous methods of removing the cation channel protein from the solid phase resin are known to those of ordinary skill in the art. In a preferred embodiment, wherein the solid phase resin is a cobalt resin, the removing step comprises contacting the cation channel protein conjugated to the solid phase resin with an imidazole solution. This solution readily cleaves any bonds conjugating the cation channel protein to the resin, so that the protein can removed from the resin, and collected for further analysis to determine whether the potential drug or agent is bound to the protein.

Problems solved by technology

Although potassium channel proteins are involved in such a wide variety of homeostatic functions, few drugs or therapeutic agents are available that act on potassium channel proteins to treat abnormal processes.

A reason for a lack of presently available drugs that act on potassium channel proteins is that isolated potassium channel proteins are not available in great abundance, mainly because an animal cell requires only a very limited number of such channel proteins in order to function.

Consequently, it has been very difficult to isolate and purify potassium channel proteins, reducing the amount of drug screening efforts in search of potassium channel protein acting drugs.

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