Control over potassium (K+) levels is essential to normal tissue homeostasis and a number of physiological processes such as electrolyte balance, cell signaling and cellular electrical activity. Potassium channels mediate the flux of potassium across the cell membrane in a highly selective manner. It is essential that channels be able to regulate the diffusion of ions, and so must be able to switch between a conductive and non-conductive state, a process known as gating. During gating, the channel’s pore width is thought to change, with an ‘open’ channel having a wide pore aperture, and a ‘closed’ channel a narrow one. This study set out to test the gating hypothesis. The conformation of a KIRK+ channel was covalently constrained at specific positions by sulfhydryl-reactive crosslinkers. Crosslinking to completion was verified, the conformation of the pore in crosslinked channels elucidated and function assessed by fluorimetric liposomal flux assays.