News and Publications

Ion Channels

N. Unwin

The goal of research in this laboratory is to understand how ion channels work by analyzing their 3-dimensional structures. My colleagues and I use cryo-electron microscopy combined with rapid freezing to trap different conformational states. Our studies center on the nicotinic acetylcholine receptor obtained from Torpedo electric organ and on voltage-gated potassium channels obtained by overexpression in insect cells. We wish to find out how such ion channels achieve their specific ion selectivities and transport rates and to understand, in physical terms, how the channels open, close, and desensitize in response to chemical or electrical stimuli.

The resolution attained so far with the nicotinic acetylcholine receptor (9 Å) is sufficient to show some elements of secondary structure. A group of 3 rods is visualized in the extracellular part of each subunit, 30--40 Å above the surface of the membrane. The 2 -subunits have a distinctive appearance in this region, suggesting that the rods may be involved in forming the binding pockets for acetylcholine. Another rod is visualized in the membrane-spanning part of each subunit, which forms the wall lining the pore. Features of these rods, which presumably are -helices, provide insight into how the binding site is designed and how the gate of the channel is constructed. Freeze-trapping experiments indicate that binding of acetylcholine to open the channel causes a localized disturbance in the extracellular domain and initiates small rotations of the protein subunits. These rotations trigger a change in the configuration of the -helical rods lining the membrane-spanning pore: the pore opens up to let the ions through.

Recently, we developed a method that allows accurate determination and 3-dimensional correction of distortions present in the acetylcholine receptor crystals. Images much better than those obtainable with standard instruments can also be obtained by using a high-voltage, field-emission microscope that incorporates a liquid helium--cooled stage developed by Y. Fujiyoshi, Kyoto, Japan. Thus, it should soon be possible to see the different conformations of the acetylcholine receptor at near-atomic resolution. Research on potassium channels is at the initial stage of exploring ways of making specimens suitable for electron crystallographic analysis.

PUBLICATIONS

Beroukhim, R., Unwin, N. Distortion correction of tubular crystals: Improvements in the acetylcholine receptor structure. Ultramicroscopy 70:57, 1997.

Unwin, N. The nicotinic acetylcholine receptor of the Torpedo electric ray. J. Struct. Biol. 121:181, 1998.