Abstract
Interconversion between conductive and non-conductive forms of the K+ channel selectivity filter underlies a variety of gating events, from flicker transitions (at the microsecond timescale) to C-type inactivation (millisecond to second timescale). Here we report the crystal structure of the Streptomyces lividans K+ channel KcsA in its open-inactivated conformation and investigate the mechanism of C-type inactivation gating at the selectivity filter from channels ‘trapped’ in a series of partially open conformations. Five conformer classes were identified with openings ranging from 12 Å in closed KcsA (Cα–Cα distances at Thr 112) to 32 Å when fully open. They revealed a remarkable correlation between the degree of gate opening and the conformation and ion occupancy of the selectivity filter. We show that a gradual filter backbone reorientation leads first to a loss of the S2 ion binding site and a subsequent loss of the S3 binding site, presumably abrogating ion conduction. These structures indicate a molecular basis for C-type inactivation in K+ channels.
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Protein Data Bank
Data deposits
The atomic coordinates and structure factors of KcsA-OM in the open-inactivated conformation and four additional partially open structural classes have been deposited with the Protein Data Bank under the following accession numbers: 3F5W; 3F7V; 3F7Y; 3FB6; and 3BF5.
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Acknowledgements
We thank F. Bezanilla, B. Roux, H. Mchaourab and R. Nakamoto for discussions and comments on the manuscript. K. Locher for comments on the manuscripts. R. Mackinnon provided the Fab-expressing hybridoma cells. We also thank S. Chakrapani, J. Cordero-Morales, J. Santos, S. Uysal, O. Dalmas and the members of the Perozo laboratory for discussions and comments on the manuscript. We are thankful to K. R. Rajashankar, R. Sanishvili and the staff at the NE-CAT 24ID and GM-CA 23ID beamlines at the Advanced Photon Source, Argonne National Laboratory. This work was supported in part by NIH grant R01-GM57846 and by a gift from the Palmer family.
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E.P. and L.G.C. conceived the project. L.G.C. and D.M.C. generated constructs, performed biochemical analysis, expressed, purified and crystallized the proteins. L.G.C. performed electrophysiology experiments. L.G.C., V.J. and E.P. collected X-ray diffraction data. L.G.C. and V.J. determined and analysed the structures. E.P., L.G.C. and V.J. analysed the data and wrote the paper.
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Supplementary information
Supplementary Information
This file contains Supplementary Table 1, Supplementary Figures S1-S10 and References. (PDF 3578 kb)
Supplementary Movie 1
This movie shows selectivity filter gating, top view. (MPG 2464 kb)
Supplementary Movie 2
This movie shows channel gating, bottom view. (MPG 4364 kb)
Supplementary Movie 3
This movie shows channel gating, side view. (MPG 2325 kb)
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Cuello, L., Jogini, V., Cortes, D. et al. Structural mechanism of C-type inactivation in K+ channels. Nature 466, 203–208 (2010). https://doi.org/10.1038/nature09153
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DOI: https://doi.org/10.1038/nature09153
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