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The conserved phenylalanine in the K+ channel voltage-sensor domain creates a barrier with unidirectional effects
KTH, School of Engineering Sciences (SCI), Theoretical Physics, Theoretical & Computational Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
Linköping University.
KTH, School of Engineering Sciences (SCI), Theoretical Physics, Theoretical & Computational Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0002-2734-2794
2013 (English)In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 104, no 1, 75-84 p.Article in journal (Refereed) Published
Abstract [en]

Voltage-gated ion channels are crucial for regulation of electric activity of excitable tissues such as nerve cells, and play important roles in many diseases. During activation, the charged S4 segment in the voltage sensor domain translates across a hydrophobic core forming a barrier for the gating charges. This barrier is critical for channel function, and a conserved phenylalanine in segment S2 has previously been identified to be highly sensitive to substitutions. Here, we have studied the kinetics of Kv1-type potassium channels (Shaker and Kv1.2/2.1 chimera) through site-directed mutagenesis, electrophysiology, and molecular simulations. The F290L mutation in Shaker (F233L in Kv1.2/2.1) accelerates channel closure by at least a factor 50, although opening is unaffected. Free energy profiles with the hydrophobic neighbors of F233 mutated to alanine indicate that the open state with the fourth arginine in S4 above the hydrophobic core is destabilized by ∼17 kJ/mol compared to the first closed intermediate. This significantly lowers the barrier of the first deactivation step, although the last step of activation is unaffected. Simulations of wild-type F233 show that the phenyl ring always rotates toward the extracellular side both for activation and deactivation, which appears to help stabilize a well-defined open state.

Place, publisher, year, edition, pages
2013. Vol. 104, no 1, 75-84 p.
Keyword [en]
Potassium Channel, Gating Charge, Transition, Kv1.2, State, Conformation, Hanatoxin, Segment, Field, S4
National Category
Biochemistry and Molecular Biology
URN: urn:nbn:se:kth:diva-104739DOI: 10.1016/j.bpj.2012.11.3827ISI: 000313541200010ScopusID: 2-s2.0-84872151920OAI: diva2:566861
Science for Life Laboratory - a national resource center for high-throughput molecular bioscienceEU, European Research Council, 209825Swedish Foundation for Strategic Research Swedish Research CouncilSwedish e‐Science Research Center

QC 20130212. Updated from manuscript to article in journal.

Available from: 2012-11-09 Created: 2012-11-09 Last updated: 2013-02-14Bibliographically approved
In thesis
1. Voltage sensor activation and modulation in ion channels
Open this publication in new window or tab >>Voltage sensor activation and modulation in ion channels
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Voltage-gated ion channels play fundamental roles in neural excitability, they are for instance responsible for every single heart beat in our bodies, and dysfunctional channels cause disease that can be even lethal. Understanding how the voltage sensor of these channels function is critical for drug design of compounds targeting neuronal excitability.

The opening and closing of the pore in voltage-gated potassium (Kv) channels is caused by the arginine-rich S4 helix of the voltage sensor domain (VSD) moving in response to an external potential. In fact, VSDs are remarkably efficient at turning membrane potential into conformational changes, which likely makes them the smallest existing biological engines. Exactly how this is accomplished is not yet fully known and an area of hot debate, especially due to the lack of structures of the resting and intermediate states along the activation pathway. In this thesis I study how the VSD activation works and show how toxic compounds modulate channel gating through direct interaction with these quite unexplored drug targets.

First, I show that a secondary structure transition from alpha- to 3(10)-helix in the S4 helix is an important part of the gating as this helix type is significantly more favorable compared to the -helix in terms of a lower free energy barrier. Second, I present new models for intermediate states along the whole voltage sensor cycle from closed to open and suggest a new gating model for S4, where it moves as a sliding 3(10)-helix. Interestingly, this 3(10)-helix is formed in the region of the single most conserved residue in Kv channels, the phenylalanine F233. Located in the hydrophobic core, it directly faces S4 and creates a structural barrier for the gating charges. Substituting this residue alters the deactivation free energy barrier and can either facilitate the relaxation of the voltage sensor or increase the free energy barrier, depending on the size of the mutant. These results are confirmed by new experimental data that supports that a rigid ring at the phenylalanine position is the rate-limiting factor for the deactivation gating process, while the activation is unaffected. Finally, we study how the activation can be modulated for pharmaceutical reasons. Neurotoxins such as hanatoxin and stromatoxin push S3b towards S4 helix limiting S4's flexibility. This makes it harder for the VSD to activate and might explain the stronger binding affinities in resting state.

All these results are highly important both for the general topic of biological macromolecules undergoing functionally critical conformational transitions, as well as the particular case of voltage-gated ion channels where understanding of the gating process is probably the key step to explain the effects of mutations or drug interactions.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2012. xiii, 77 p.
Trita-FYS, ISSN 0280-316X ; 2012:77
activation, deactivation, inactivation, voltage sensor, VSD, gating, Kv1.2-2.1, Shaker, F233, hydrophobic barrier
National Category
Biological Sciences
urn:nbn:se:kth:diva-104742 (URN)978-91-7501-498-2 (ISBN)
Public defence
2012-12-07, FB53, AlbaNova universitetscentrum, Stockholm, 10:00 (English)
Swedish e‐Science Research CenterScience for Life Laboratory - a national resource center for high-throughput molecular bioscience

QC 20121115

Available from: 2012-11-15 Created: 2012-11-09 Last updated: 2013-04-15Bibliographically approved

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