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Molecular driving forces determining potassium channel slow inactivation

Abstract

K+ channels undergo a time-dependent slow inactivation process that plays a key role in modulating cellular excitability. Here we show that in the prokaryotic proton-gated K+ channel KcsA, the number and strength of hydrogen bonds between residues in the selectivity filter and its adjacent pore helix determine the rate and extent of C-type inactivation. Upon channel activation, the interaction between residues at positions Glu71 and Asp80 promotes filter constriction parallel to the permeation pathway, which affects K+-binding sites and presumably abrogates ion conduction. Coupling between these two positions results in a quantitative correlation between their interaction strength and the stability of the inactivated state. Engineering of these interactions in the eukaryotic voltage-dependent K+ channel Kv1.2 suggests that a similar mechanistic principle applies to other K+ channels. These observations provide a plausible physical framework for understanding C-type inactivation in K+ channels.

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Figure 1: The Glu71-Asp80 interaction is the driving force for inactivation in KcsA.
Figure 2: Inverse relationship between the rate of inactivation and the open dwell times.
Figure 3: Histidine at position 71 inverts the voltage dependence of inactivation.
Figure 4: Structural basis of inactivation in E71S and E71H channels.
Figure 5: Relation between the open-channel probability (evaluated from Gaussian fits of histograms of all time points) and equilibrium X71-Asp80 Cα–Cα distance extracted from molecular dynamics simulations and crystal structures.
Figure 6: Correlation between the extent of inactivation and the energetics of filter distortion.
Figure 7: Mutations at position 370 recapitulate C-type inactivation in Kv1.2.
Figure 8: Structural rearrangement during C-type inactivation at the filter.

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Acknowledgements

We thank F. Bezanilla, S. Chakrapani, L. Cuello, M. Sotomayor and H. Raghuraman for critical reading and discussion of the manuscript; M. Wiener and M. Purdy for crystallographic data collection (for E71T); J. Faraldo-Gomez and A. Lau for critical comments on the PMF calculations; S. Goldstein (University of Chicago) for providing access to the two electrode voltage clamp system; the staff of the GM/CA-23-ID beamline at the Advanced Photon Source for their invaluable assistance in data collection; R. MacKinnon (Rockefeller University) for providing the KcsA antibody hybridoma cell line; and the National Center for Supercomputing Applications and Jazz computing cluster at Argonne National Laboratory for computer time. This work was supported by US National Institutes of Health grants to E.P. and B.R.

Author information

Authors and Affiliations

Authors

Contributions

J.F.C.-M. carried out the channel mutagenesis, biochemistry, EPR studies, crystallization and electrophysiology with KcsA channels, and J.F.C.-M. and A.L. carried these out on Kv1.2 channels. V.J. carried out crystal data collection, structure solving and computational analyses. V.V. participated in channel mutagenesis, biochemistry and EPR. D.M.C. made all the Fab preparations. B.R., working with V.J., participated in computational design and PMF calculations. E.P. directed experimental design and data analyses, and wrote the manuscript with J.F.C.-M., V.J. and B.R.

Corresponding author

Correspondence to Eduardo Perozo.

Supplementary information

Supplementary Text and Figures

Supplementary Figure 1–6, Supplementary Table 1 (PDF 4408 kb)

Supplementary Video 1

E71H movie. These movies were made with VMD. VMD is developed with NIH support by the Theoretical and Computational Biophysics group at the Beckman Institute, University of Illinois at Urbana-Champaign. (http://www.ks.uiuc.edu/)7. All the molecular graphic figures in this work were made using Pymol (DeLano, W.L. The PyMOL Molecular Graphics System (2002) http://pymol.sourceforge.net/). (MPG 3664 kb)

Supplementary Video 2

E71A movie. These movies were made with VMD. VMD is developed with NIH support by the Theoretical and Computational Biophysics group at the Beckman Institute, University of Illinois at Urbana-Champaign. (http://www.ks.uiuc.edu/)7. All the molecular graphic figures in this work were made using Pymol (DeLano, W.L. The PyMOL Molecular Graphics System (2002) http://pymol.sourceforge.net/). (MPG 3445 kb)

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Cordero-Morales, J., Jogini, V., Lewis, A. et al. Molecular driving forces determining potassium channel slow inactivation. Nat Struct Mol Biol 14, 1062–1069 (2007). https://doi.org/10.1038/nsmb1309

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