Potassium Ion Channel

by: Jim_Kling


X-ray crystallography is a vital tool for studying the structure and function of proteins. It relies on coaxing a protein into forming a well-ordered crystal—a demanding task that can take months or even years to accomplish. To decipher the structure, researchers bombard the crystal with X-rays to produce a diffraction pattern that can be decoded to produce a three-dimensional picture of an individual protein.
However, many proteins can adopt multiple structures, so a natural question arises: Is the structure a protein adopts in a crystal identical to the one it adopts in its cellular environment? This question lies at the center of a debate over the structure of KvAP, a voltage-dependent potassium ion channel taken from a bacterium found in ocean bottom hydrothermal vents. In December, Science magazine named a group of journal articles about the issue one of its top 10 Science breakthroughs of 2005.
In two articles in Nature in 2003, a team led by Roderick MacKinnon of Rockefeller University published two structures of KvAP Such channels are vital to electrical impulses’ conduction along neurons and other cells. The proteins are embedded in cell membranes, where they sense changes in voltage potential across the membrane, opening or closing the pore to regulate the cell's potassium ion content and adjust the voltage potential.
The protein was particularly challenging to crystallize because it exists within the cell membrane, which places it in a hydrophobic environment and also physically constrains it. To accomplish the task, the researchers attached antibody fragments to the protein to stabilize it. They produced two crystal structures—one of the entire protein and one of just the regions that act as voltage sensors. The resulting structures revealed a central ion conduction pore surrounded by four paddle-like voltage sensors that spread out from the channel's outer perimeter. The protein also contains flexible hinges. From this structure, the researchers concluded that charged amino acids in the voltage sensor 'paddles' cause them to move in response to a change in the voltage across the membrane. The hinges transfer that movement to the pore, causing it to open or close.
The structures provided evidence for a novel mechanism for voltage-dependent potassium ion channels, but they also seemed slightly askew. The paddle-like voltage sensors were tilted away from the direction that many had expected based on the physiological measurements of the potassium ion channel. The researchers concluded that the structure was a 'non-native' conformation. Did the use of antibody fragments somehow alter the structure of the protein in the crystal?
In the October 25, 2005 edition of the Proceedings of the National Academy of Sciences, the team published a follow-up study that answered that question. In this work, they produced two additional crystal structures of KvAP, one using different antibody fragments than those used in the earlier crystallization, and one without using any antibody fragments at all. Both had similar structures to the original, indicating that the antibody fragments were not responsible for the non-native conformation.
In fact, the researchers suggest that the change is a result of removing the protein from the cell membrane. The voltage sensors are not tightly linked to the pore, but the cell membrane helps lock them into position. When the protein leaves that environment in preparation for crystallization, that constraint is removed, allowing the protein to reorient itself.
The nature of KvAP probably contributed to the difficulty in getting a native conformation in the crystal structure. For the voltage sensors to cause the opening and closing of the ion pore, the protein must be highly flexible, which in turn makes it capable of adopting a number of different conformations. In fact, it likely has a number of native structures, MacKinnon says. There may be many more yet to discover.

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This article first appeared on February 6, 2006. From: http://www.chemistry.org/portal/a/c/s/1/feature_pro.html?id=c373e9092cbda5d78f6a17245d830100