Charybdotoxin unbinding from the mKv1.3 potassium channel: A combined computational and experimental study

Khabiri M¹, Nikouee A², Grissmer S², Ettrich R¹

¹Institute of Nanobiology and Structural Biology of GCRC, AS CR, and Faculty of Sciences, University of South Bohemia in České Budějovice, 37333 Nove Hrady, Czech Republic

²Institute of Applied Physiology, Ulm University, Ulm, Germany

Charybdotoxin, belonging to the group of so-called scorpion toxins, is a short peptide able to block many voltage-gated potassium channels, such as mKv1.3, with high affinity. We use a reliable homology model based on the high-resolution crystal structure of the 94% sequence identical homolog Kv1.2 for charybdotoxin docking followed by molecular dynamics simulations to investigate the mechanism and energetics of unbinding, tracing the behavior of the channel protein and charybdotoxin during umbrella-sampling simulations as charybdotoxin is moved away from the binding site. The potential of mean force is constructed from the umbrella sampling simulations and combined with K_d and free energy values gained experimentally using the patch-clamp technique to study the free energy of binding at different ion concentrations and the mechanism of the charybdotoxin-mKv1.3 binding process. A possible charybdotoxin binding mechanism is deduced that includes an initial hydrophobic contact followed by stepwise electrostatic interactions and finally optimization of hydrogen-bonds and salt-bridges.

[1] Morteza Khabiri, Azadeh Nikouee, Lukasz Cwiklik, Stephan Grissmer, Rüdiger Ettrich (2011) Charybdotoxin unbinding from the mKv1.3 potassium channel: A combinedcomputational and experimental study Journal of Physical Chemistry B 115: 39. 11490–11500.

[2] Azadeh Nikouee, Morteza Khabiri, Stephan Grissmer, Rüdiger Ettrich (2012) Charybdotoxin and margatoxin acting on the human voltage-gated potassium channel hKv1.3 and itsH399N mutant: An experimental and computational comparison Journal of Physical Chemistry B, accepted for revision 21.12.2011

M.K. and R.E. acknowledge support from the Czech Science Foundation, Grant 203/08/0114, and the University of South Bohemia, Grant GAJU 170/2010/P. S.G. and A.N. were supported by grants from the DFG (Gr 848/14-1). Access to the METACentrum computing facilities provided under the research intent MSM6383917201 is highly appreciated.