Dokument

Zusammenfassung:
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 (Kv-Kanäle) und blockiert selektiv Kv1-Ionenkanäle. In der hier vorliegenden Arbeit wurde mittels zielgerichteter Mutagenese, Alanin-Scans, Dockings, molekulardynamischen Simulationen sowie „inside-out macropatch“-Experimenten gezeigt, dass Psora-4 in der hochkonservierten Bindungsstelle in der zentralen Kavität bindet. Zusätzlich bindet Psora-4 in vier weniger konservierten hydrophoben Seitentaschen, die sich auf der „Rückseite“ der porenformenden S5- und S6-Segmente einer Untereinheit und dem Spannungssensor S4 und dem S4-S5-Linker der benachbarten Untereinheit befinden. Das gleichzeitige Binden an beiden Bindungsstellen führt zur Bildung eines stabilen, nicht-leitenden Inhibitor-Ionenkanal-Komplexes. Diese Studie zeigt einen neuen Blockmechanismus, der die hohe Affinität, die Kooperativität, das benötigte Öffnen und Schließen des Ionenkanals (“use dependence“) und die Selektivität für Kv1-Ionenkanäle erklärt. Die neu identifizierte, weniger konservierte Bindungsstelle in den hydrophoben Seitentaschen stellt die molekulare Basis für die Entwicklung spezifischer Kv-Kanal-Blocker dar. Die Inaktivierungsrate der Kv-Kanäle ist ein entscheidender Faktor zur Frequenzmodulation in Neuronen. Mehrfach ungesättige Fettsäuren (Polyunsaturated fatty acids: PUFAs) modulieren die Kinetik von Kv-Kanälen und beeinflussen so die neuronale und kardiale Erregungsweiterleitung. Diese Studie zeigt, dass PUFAs an der Bindungsstelle in der zentralen Kavität binden und so die Passage von Ionen verhindern. Außerdem ergibt ein systematischer Alanin-Scan, der 132 Aminosäuren im S3-S4-Linker, S4-Segment, S4-S5-Linker, S5-Segment, den Porenschleifen und dem S6-Segment umfasst, ein deutlich erweitertes Bild der Interaktion zwischen PUFAs und Kv-Kanälen. Diese Studie zeigt, dass zusätzlich zur Blockade der zentralen Pore auch die Porenschleifen, die hydrophoben Seitentaschen, die lateralen Fenster und der Spannungssensor an der Modulation der Kinetik der PUFA-Inhibition beteiligt sind.

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