Dataset Information

A limited access compartment between the pore domain and cytosolic domain of the BK channel.

ABSTRACT: Cytosolic N-terminal segments of many K+ channel subunits mediate rapid blockade of ion permeation by physical occlusion of the ion-conducting pore. For some channels with large cytosolic structures, access to the channel pore by inactivation domains may occur through lateral entry pathways or "side portals" that separate the pore domain and associated cytosolic structures covering the axis of the permeation pathway. However, the extent to which side portals control access of molecules to the channel or influence channel gating is unknown. Here we use removal of inactivation by trypsin as a tool to examine basic residue accessibility in both the N terminus of the native auxiliary beta2 subunit of Ca2+-activated, BK-type K+ channels and beta2 subunits with artificial inactivating N termini. The results show that, for BK channels, side portals define a protected space that precedes the channel permeation pathway and excludes small proteins such as trypsin but allows inactivation domains to enter. When channels are closed, inactivation domains readily pass through side portals, with a central antechamber preceding the permeation pathway occupied by an inactivation domain approximately half of the time under resting conditions. The restricted volume of the pathway through side portals is likely to influence kinetic properties of inactivation mechanisms, blockade by large pharmacological probes, and accessibility of modulatory factors to surfaces of the channel within the protected space.

SUBMITTER: Zhang Z

PROVIDER: S-EPMC6674862 | biostudies-literature | 2006 Nov

REPOSITORIES: biostudies-literature

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A limited access compartment between the pore domain and cytosolic domain of the BK channel.

Zhang Zhe Z Zhou Yu Y Ding Jiu-Ping JP Xia Xiao-Ming XM Lingle Christopher J CJ

The Journal of neuroscience : the official journal of the Society for Neuroscience 20061101 46

Cytosolic N-terminal segments of many K+ channel subunits mediate rapid blockade of ion permeation by physical occlusion of the ion-conducting pore. For some channels with large cytosolic structures, access to the channel pore by inactivation domains may occur through lateral entry pathways or "side portals" that separate the pore domain and associated cytosolic structures covering the axis of the permeation pathway. However, the extent to which side portals control access of molecules to the ch ...[more]

PMID: 17108156

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Project description:The internal vestibule of large-conductance Ca(2+) voltage-activated K(+) (BK) channels contains a ring of eight negative charges not present in K(+) channels of lower conductance (Glu386 and Glu389 in hSlo) that modulates channel conductance through an electrostatic mechanism (Brelidze, T.I., X. Niu, and K.L. Magleby. 2003. Proc. Natl. Acad. Sci. USA. 100:9017-9022). In BK channels there are also two acidic amino acid residues in an extracellular loop (Asp326 and Glu329 in hSlo). To determine the electrostatic influence of these charges on channel conductance, we expressed wild-type BK channels and mutants E386N/E389N, D326N, E329Q, and D326N/E329Q channels on Xenopus laevis oocytes, and measured the expressed currents under patch clamp. Contribution of E329 to the conductance is negligible and single channel conductance of D326N/E329Q channels measured at 0 mV in symmetrical 110 mM K(+) was 18% lower than the control. Current-voltage curves displayed weak outward rectification for D326N and the double mutant. The conductance differences between the mutants and wild-type BK were caused by an electrostatic effect since they were enhanced at low K(+) (30 mM) and vanished at high K(+) (1 M K(+)). We determine the electrostatic potential change, Deltaphi, caused by the charge neutralization using TEA(+) block for the extracellular charges and Ba(2+) for intracellular charges. We measured 13 +/- 2 mV for Deltaphi at the TEA(+) site when turning off the extracellular charges, and 17 +/- 2 mV for the Deltaphi at the Ba(2+) site when the intracellular charges were turned off. To understand the electrostatic effect of charge neutralizations, we determined Deltaphi using a BK channel molecular model embedded in a lipid bilayer and solving the Poisson-Boltzmann equation. The model explains the experimental results adequately and, in particular, gives an economical explanation to the differential effect on the conductance of the neutralization of charges D326 and E329.

Dataset Information

A limited access compartment between the pore domain and cytosolic domain of the BK channel.

Publications

A limited access compartment between the pore domain and cytosolic domain of the BK channel.

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