NMR Structure of the “Ball-and-chain” Domain of KCNMB2, the β2-Subunit of Large Conductance Ca2+- and Voltage-activated Potassium Channels

Bentrop, Detlef; Beyermann, Michael; Wissmann, Ralph; Fakler, Bernd

doi:10.1074/jbc.m107118200

Cited by 61 publications

(62 citation statements)

References 34 publications

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“…For channels in resting states, relative differences in digestion rates of particular basic residues will primarily reflect intrinsic structural characteristics of the ␤2 N terminus. In this regard, the present results are, in part, consistent with nuclear magnetic resonance (NMR) structures previously reported for the isolated 45 amino acid N-terminal ␤2 peptide (Bentrop et al, 2001). The isolated peptide in solution adopts a large family of structures, with two ␣-helical segments, residues 10 -17 and 20 -30.…”

Section: Discussionsupporting

confidence: 80%

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A Limited Access Compartment between the Pore Domain and Cytosolic Domain of the BK Channel

Zhang¹,

Zhong²,

Ding³

et al. 2006

J. Neurosci.

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Cytosolic N-terminal segments of many K ϩ 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 ␤2 subunit of Ca 2ϩ -activated, BK-type K ϩ channels and ␤2 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.

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Section: Discussionsupporting

confidence: 80%

“…Comparison of the structures of ␤2 (1JO6) (Bentrop et al, 2001), MthK (1LNQ) (Jiang et al, 2002a), and porcine pancreatic trypsin A-chain (1AVW) (Song and Suh, 1998) was done with VMD and PyMOL (DeLano, 2002).…”

Section: Modeling Of the Trypsin-mediated Removal Of Inactivationmentioning

confidence: 99%

A Limited Access Compartment between the Pore Domain and Cytosolic Domain of the BK Channel

Zhang¹,

Zhong²,

Ding³

et al. 2006

J. Neurosci.

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“…The 15 N relaxation measurements (T 1 , T 2 , 1 H-15 N NOE) were carried out at 500-MHz proton frequency as described previously (23) with overall delays between scans of 2 s for the T 1 and T 2 measurements and 3 s for the heteronuclear NOE experiment, respectively. The delays used for the T 1 experiments were 10, 20, 40, 80, 160, 320, 640, 1280, and 2560 ms, whereas delays of 8,16,24,40,64,96,128,160,256,320,480, and 800 ms were used for the T 2 experiments. The 15 N relaxation data were analyzed according to the reduced spectral density mapping approach (24).…”

Section: Methodsmentioning

confidence: 99%

“…Instead, the rate of association between ID and receptor must be decreased. In the "sequential-step" model of inactivation (13,17,40), such association actually implies two separate steps: first, the approach of the ID toward the channel entrance (preinactivation) and, second, the binding of the ID to its receptor (inactivation). In this context, the alanine-rich helix in ID2 may be expected to work as a "docking domain" that binds the N terminus to the surface of the channel vestibule and thus promotes rapid insertion of ID1 into its receptor, which finally occludes the pore and inactivates the channel (see also "Discussion").…”

Section: Structure-function Analysis Delineates Distinct Properties Amentioning

confidence: 99%

Solution Structure and Function of the “Tandem Inactivation Domain” of the Neuronal A-type Potassium Channel Kv1.4

Wissmann

Bildl

Oliver

et al. 2003

Journal of Biological Chemistry

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Cumulative inactivation of voltage-gated (Kv) K؉ channels shapes the presynaptic action potential and determines timing and strength of synaptic transmission. Kv1.4 channels exhibit rapid "ball-and-chain"-type inactivation gating. Different from all other Kv␣ subunits, Kv1.4 harbors two inactivation domains at its N terminus. Here we report the solution structure and function of this "tandem inactivation domain" using NMR spectroscopy and patch clamp recordings. Inactivation domain 1 (ID1, residues 1-38) consists of a flexible N terminus anchored at a 5-turn helix, whereas ID2 (residues 40 -50) is a 2.5-turn helix made up of small hydrophobic amino acids. Functional analysis suggests that only ID1 may work as a pore-occluding ball domain, whereas ID2 most likely acts as a "docking domain" that attaches ID1 to the cytoplasmic face of the channel. Deletion of ID2 slows inactivation considerably and largely impairs cumulative inactivation. Together, the concerted action of ID1 and ID2 may promote rapid inactivation of Kv1.4 that is crucial for the channel function in short term plasticity.

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“…Based on the results from Western Blot, we deduced the molecular weight of a carbohydrate chain which is about 3,000 Da, and hypothesized the stucture of carbohydrate chain (LinucsID 3961) from glycosciences.de (http://glycosciences.de, a web databases and bioinformatics tools for glycobiology and glycomics). 31 The full model of b2 subunit was then constructed by connecting the predicted extracellular b2 loop, 2 transmembrane helices, N-terminal 32 (PDB ID: 1JO6) and the carbohydrate chains. The full models of BK channels were built as previous.…”

Section: Patch Clamp Recordingmentioning

confidence: 99%

The glycosylation of the extracellular loop ofβ2 subunits diversifies functional phenotypes of BK Channels

et al. 2016

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Large-conductance Ca 2C -and voltage-activated potassium (MaxiK or BK) channels are composed of a pore-forming a subunit (Slo) and 4 types of auxiliary b subunits or just a pore-forming a subunit. Although multiple N-linked glycosylation sites in the extracellular loop of b subunits have been identified, very little is known about how glycosylation influences the structure and function of BK channels. Using a combination of site-directed mutagenesis, western blot and patch-clamp recordings, we demonstrated that 3 sites in the extracellular loop of b2 subunit are N-glycosylated (N-X-T/S at N88, N96 and N119). Glycosylation of these sites strongly and differentially regulate gating kinetics, outward rectification, toxin sensitivity and physical association between the a and b2 subunits. We constructed a model and used molecular dynamics (MD) to simulate how the glycosylation facilitates the association of a/b2 subunits and modulates the dimension of the extracellular cavum above the pore of the channel, ultimately to modify biophysical and pharmacological properties of BK channels. Our results suggest that N-glycosylation of b2 subunits plays crucial roles in imparting functional heterogeneity of BK channels, and is potentially involved in the pathological phenotypes of carbohydrate metabolic diseases.

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NMR Structure of the “Ball-and-chain” Domain of KCNMB2, the β2-Subunit of Large Conductance Ca2+- and Voltage-activated Potassium Channels

Cited by 61 publications

References 34 publications

A Limited Access Compartment between the Pore Domain and Cytosolic Domain of the BK Channel

A Limited Access Compartment between the Pore Domain and Cytosolic Domain of the BK Channel

Solution Structure and Function of the “Tandem Inactivation Domain” of the Neuronal A-type Potassium Channel Kv1.4

The glycosylation of the extracellular loop ofβ2 subunits diversifies functional phenotypes of BK Channels

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