Computational design of water-soluble analogues of the potassium channel KcsA

doi:10.1073/pnas.0306417101

Computational design of water-soluble analogues of the potassium channel KcsA

^*Department of Biochemistry and Molecular Biophysics, Johnson Foundation, School of Medicine, ^§Makineni Theoretical Laboratories, and ^¶Department of Chemistry, University of Pennsylvania, Philadelphia, PA 19104; and ^†Neutron Science Research Center and Center for the Promotion of Computational Science and Engineering, Japan Atomic Energy Research Institute, 8-1, Umemidai, Kizu-cho, Souraku-gun, Kyoto 619-0215, Japan

Edited by Douglas C. Rees, California Institute of Technology, Pasadena, CA, and approved December 4, 2003 (received for review October 5, 2003)

Abstract

Although the interiors of membrane and water-soluble proteins are similar in their physicochemical properties, membrane proteins differ in having larger fractions of hydrophobic residues on their exteriors. Thus, it should be possible to water-solubilize membrane proteins by mutating their lipid-contacting side chains to more polar groups. Here, a computational approach was used to generate water-soluble variants of the potassium channel KcsA. As a probe of the correctness of the fold, the proteins contain an agitoxin2 binding site from a mammalian homologue of the channel. The resulting proteins express in high yield in Escherichia coli and share the intended functional and structural properties with KcsA, including secondary structure, tetrameric quaternary structure, and tight specific binding to both agitoxin2 and a small molecule channel blocker.

Footnotes

↵ ‡ To whom correspondence may be addressed. E-mail: lear{at}mail.med.upenn.edu, saven{at}sas.upenn.edu, or wdegrado{at}mail.med.upenn.edu.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: AgTx₂, agitoxin2; TEA, tetraethylammonium chloride; AgTx₂-DNP, AgTx₂ with 2,4-dinitrophenyl-Ala; E _env, environmental energy.