Abstract
C-type inactivation in potassium channels helps fine-tune long-term channel activity through conformational changes at the selectivity filter. Here, through the use of cross-linked constitutively open constructs, we determined the structures of KcsA's mutants that stabilize the selectivity filter in its conductive (E71A, at 2.25 Å) and deep C-type inactivated (Y82A at 2.4 Å) conformations. These structural snapshots represent KcsA's transient open-conductive (O/O) and the stable open deep C-type inactivated states (O/I), respectively. The present structures provide an unprecedented view of the selectivity filter backbone in its collapsed deep C-type inactivated conformation, highlighting the close interactions with structural waters and the local allosteric interactions that couple activation and inactivation gating. Together with the structures associated with the closed-inactivated state (C/I) and in the well-known closed conductive state (C/O), this work recapitulates, at atomic resolution, the key conformational changes of a potassium channel pore domain as it progresses along its gating cycle.
Highlights
The simplest description of the gating cycle in the pore domain of a K+ channel requires at least four distinct kinetic states (Ostmeyer et al, 2013; Panyi and Deutsch, 2006; Yellen, 1998)
As a consequence of allosteric coupling between the inner gate and the pore helix near the selectivity filter (Cuello et al, 2010a; Pan et al, 2011), opening of the activation gate in many channels leads to a series of structural changes at the selectivity filter that renders it non-conductive, a process known as C-type inactivation (Hoshi et al, 1991; Liu et al, 1996; Lopez-Barneo et al, 1993)
Using the open-inactivated structure of KcsA (PDB = 3F5W) (Cuello et al, 2010b) as a reference (Figure 1b), six cysteine pairs were tested between the N-terminal side of the transmembrane segment 1 (TM1) and the C-terminal end of TM2 (Figure 1c) for disulfide-bond formation in SDS gels (Figure 1d)
Summary
The simplest description of the gating cycle in the pore domain of a K+ channel requires at least four distinct kinetic states (Ostmeyer et al, 2013; Panyi and Deutsch, 2006; Yellen, 1998). The pore domain’s activation gate formed by the inner helix bundle is closed (C) while the selectivity filter is presumably conductive (O). We define this conformation as the C/O state. When an activating stimulus drives the opening of the channel’s inner helix bundle gate, the selective flow of K+ can be sustained for hundredths of milliseconds through the open (O/O) state. As a consequence of allosteric coupling between the inner gate and the pore helix near the selectivity filter (Cuello et al, 2010a; Pan et al, 2011), opening of the activation gate in many channels leads to a series of structural changes at the selectivity filter that renders it non-conductive (the O/I state), a process known as C-type inactivation (Hoshi et al, 1991; Liu et al, 1996; Lopez-Barneo et al, 1993). Allosteric interactions with the activation gate reset the selectivity filter back to its conductive conformation (C/O), completing the gating cycle (Figure 1a)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
