Die molekulare Pharmakologie der Kv1-Ionenkanalfamilie - systematische Analyse und Charakterisierung der Modulation durch Psora-4 und Arachidonsäure

Die klassische Medikamenten-Bindungsstelle für kleine, hydrophobe Inhibitoren in spannungsgesteuerten Ionenkanälen befindet sich in der hochkonservierten zentralen Kavität. Überraschenderweise unterscheidet der Inhibitor Psora-4 zwischen verschiedenen Subtypen der spannungsgesteuerten Kalium-Kanäle...

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Bibliographic Details
Main Author: Marzian, Stefanie
Contributors: Decher, Niels (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2015
Online Access:PDF Full Text
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The classical drug binding site for small molecule inhibitors of voltage-gated ion channels is located in the highly conserved central cavity. Surprisingly the inhibitor Psora-4 discriminates between different voltage-gated potassium channel (Kv channel) subtypes and selectively blocks Kv1 channels. Ala-scanning mutagenesis, molecular modeling and inside-out macropatch-experiments show that Psora-4 binds to the highly conserved drug binding site in the central cavity and additionally to four less conserved hydrophobic binding pockets located on the “backsides” of the pore forming S5 and S6 and the pore helix of one subunit and the voltage sensor S4 and the S4-S5 linker of the neighboring subunit. Simultaneous occupation of both binding sites results in a stable non-conducting drug-channel complex. Thus, this study reveals a novel blocking mechanism which confers high affinity, cooperativity, use-dependence and selectivity to Psora-4 inhibition of Kv1 channels. The newly identified, less conserved binding site in the hydrophobic side pockets provides a molecular basis to develop specific Kv channel blockers. The rate of inactivation of Kv channels is a critical factor of the neuronal firing rate. Polyunsaturated fatty acids (PUFAs) modulate the kinetics of Kv channels and thus influence neuronal and cardiac signaling. This study reveals that PUFAs bind to the drug binding site in the central cavity and occlude the permeation pathway. Furthermore, systematic ala-scanning mutagenesis including 132 residues in S3-S4 linker, S4, S4-S5 linker, S5, pore loops and S6 provides an advanced view of the interaction between PUFAs and Kv channels. This study reports that in addition to the pore block the pore loops, the hydrophobic side pockets, lateral fenestrations and the voltage sensor contribute to the modulation of PUFA blocking kinetics.