Opening Potassium Channels to Scrutiny

Crystal structure of "open" K+ channel leads to new ideas on how it works.

Nov 1, 2007
David Secko

Voltage-gated potassium (Kv) channels play an important role in modulating the electrical activity of cells, opening in response to changes in membrane voltage and allowing potassium ions to escape. Hodgkin and Huxley laid out a model in 1952 for what voltage-gated channels might be doing, but their structure remained a mystery for decades. In 2003, Roderick MacKinnon's group at Rockefeller University solved one structure of a bacterial Kv homolog, KvAP,1 but the results didn't fit predictions from previous experiments. "The world stood in consternation because the structure didn't look at all like what people had anticipated," says Benoit Roux from the University of Chicago, who was not involved in MacKinnon's findings.

In the back-to-back Hot Papers featured here, MacKinnon's group returned in 2005 with additional work on the structure of Kv channels, first reporting the crystalline structure of an apparently open mammalian Kv channel, Kv1.2.2 This was the first time the structure of an entire mammalian Kv channel had been seen. In the second paper, the team used the structure to explore how Kv1.2 senses cell membrane voltage.3 This time around, the Kv1.2 structure fit past results more neatly, and it's now sparking efforts to see the channel's other states.

Opening Up Kv Structure

Kv channels have three essential parts: the pore, the gate, and the voltage sensor. The voltage sensor detects membrane voltage, and with a change in conformation - involving a key helix called S4 - it opens the gate. The loose protein structure of Kv channels makes getting an atomic-level picture of its structure difficult, since many channels will not form usable crystals. After five years of work, MacKinnon's group coaxed KvAP to form crystals by binding it to an antibody.1 But to many "this structure was distorted," says Richard Horn (who was not involved in the study) from Thomas Jefferson University in Philadelphia, leading to debate and a flurry of research on Kv channels.

<figcaption>Using novel techniques, Long et al. crystallized the Kv1.2 potassium channel (shown here at 2.9 Å). The pore and voltage sensors (red) are seen with its associated T1 and β domains (blue) that sit outside the membrane and whose function largely remains a mystery. Credit: COURTESY OF STEPHEN LONG</figcaption>
Using novel techniques, Long et al. crystallized the Kv1.2 potassium channel (shown here at 2.9 Å). The pore and voltage sensors (red) are seen with its associated T1 and β domains (blue) that sit outside the membrane and whose function largely remains a mystery. Credit: COURTESY OF STEPHEN LONG

Stephen Long, first author on the Kv1.2 paper and now at Memorial Sloan-Kettering Cancer Center, says he and his colleagues did away with the antibody for Kv1.2. Instead, they "used a combination of lipids and detergents," says Long, a new technique that gave them crystals reminiscent of a lipid-bilayer, as detailed in the first Hot Paper. In the second Hot Paper, the resulting structure shows the voltage sensors as upright, independent domains. This wasn't predicted before hand, but it is consistent with previous results and therefore suggests that the protein is in a native state. Horn says the structure made a lot more sense.

The idea of voltage sensors as independent domains was recently supported by David Clapham from Harvard Medical School and Yasushi Okamura from Okazaki Institute for Integrative Bioscience, who independently discovered voltage-sensor proteins without pores.4,5 These proteins conduct H+ ions and might be the long sought after voltage-gated H+ channels involved in processes such as oxidative bursts in leukocytes. Most people thought of the Kv channel structure as requiring a pore to hold the voltage sensors together, since they are highly charged yet span the membrane, says Michael Grabe from the University of Pittsburgh, who was not involved in the study. "That you can chop these proteins up, and they work, no one would have believed five years ago," says Grabe.

Closing in on the Details

Despite the insight gleaned from the Kv1.2 structure, it shows only the channel in an open state. "To understand how the voltage sensor moves from one state to another, we need to see it in the down [closed] state," says Grabe. Seeing the closed state would help solve another controversy: How far does the S4 segment move between states?

Recently, Grabe and colleagues took a closer look at the S4 segment in the closed state by using conditional lethal/second-site suppressor yeast screens.6 Their works suggests that the movement of S4 is quite large, on the order of 12 Ångstroms. Other work by Francisco Bezanilla at the University of Chicago and colleagues, using histidine scanning mutagenesis to determine the constraints of the S4 segments, suggests its movement is smaller (6.5 Å).7 "There is still some discrepancy," says Horn, "but each side is kind of morphing towards the other." Horn and others agree that a closed crystalline structure will likely appear soon.

The Kv1.2 structure also left other things unanswered, since some of the structure (such as the voltage sensors) is blurred and at low resolution, says Roux. "So there is a bit of debate about details." Roux and his colleagues recently used the Kv1.2 structure in computer simulations to model these low resolutions areas, surprisingly predicting that the membrane is thinner around the ion channel.8 This could make the electric field stronger around the channel, and voltage sensing easier.

In terms of working out the details, "it's a lot of work to basically climb this one Kv1.2 mountain," says Roux, but it's leading the way to other Kv structure determinations and aiding scientists in understanding pathologies associated with ion channels.

References

1. Y. Jiang et al., "X-ray structure of a voltage-dependent K+ channel," Nature, 423:33-41, 2003. 2. S.B. Long et al., "Crystal structure of a mammalian voltage-dependent Shaker family K+ channel," Science, 309:897-903, 2005. (Cited in 246 papers) 3. S.B. Long et al., "Voltage sensor of Kv1.2: structural basis of electromechanical coupling," Science, 309:903-8, 2005. (Cited in 123 papers) 4. I.S. Ramsey et al., "A voltage-gated proton-selective channel lacking the pore domain," Nature, 440:1213­6, 2006. 5. M. Sasaki et al., "A voltage sensor-domain protein is a voltage-gated proton channel," Science, 312:589-92, 2006. 6. M. Grabe et al., "Structure prediction for the down state of a potassium channel voltage sensor," Nature, 445:550-3, 2007. 7. F.V. Campos et al., "Two atomic constraints unambiguously position the S4 segment relative to S1 and S2 segments in the closed state of Shaker K channel," Proc Natl Acad Sci, 104:7904-9, 2007. 8. V. Jogini, B. Roux, "Dynamics of the Kv1.2 voltage-gated K+ channel in a membrane environment," Biophys J, e-pub ahead of print, Aug. 17, 2007.
Data derived from the Science Watch/Hot Papers database and the Web of Science (Thomson ISI) show that Hot Papers are cited 50 to 100 times more often than the average paper of the same type and age. S.B. Long et al., "Crystal structure of a mammalian voltage-dependent Shaker family K+ channel," Science, 309:897-903, 2005. (Cited in 246 papers) S.B. Long et al., "Voltage sensor of Kv1.2: Structural basis of electromechanical coupling," Science, 309:903-8, 2005. (Cited in 123 papers)