| Summary: | 碩士 === 國立臺灣大學 === 口腔生物科學研究所 === 90 === Voltage-gated potassium channels are found in a wide variety of tissues, where their primary role is to respond to the membrane excitation to allow the repolarization phase of an action potential to occur and therefore the K+ ions can efflux. Such channels are normally homotetramers and each subunit contains four voltage-sensing transmembrane segments, namely S1 through S4, whereas S5 and S6 form the pore. Among them, S4 may play the most crucial role in sensing the voltage change. In addition, it was generally believed that the C-terminus of S3 (S3C) interacts with gating modifier toxin, like Hanatoxin, and thus has influences on the voltage required for gating. The secondary structural arrangement of S3C has been, due to such studies, intensively analyzed and the existence of an independent a-helix was then suggested.
Kv2.1, a member of shab potassium channel family, is one of the most commonly applied channels in studying of the structural-functional correlation for voltage sensing. Upon binding of Hanatoxin 1, the midpoint of the curve for required gating potential of Kv2.1 can be shifted to the right, which means more difficult to open the channels under the same condition. On the contrary, shaker channels do not show similar effects. Due to the lack of complete structure in high resolution for S3C, the study of the structural-functional correlation have been examined only with experiments in eletrophysiology. We have designed a series of experiments to investigate the functional roles of the vicinity around S3, S4 and S3-S4 linker in affecting the gating behavior.
Regarding the stereochemistry, the electrostatic properties, as well as the hydrophobicity, and upon the utilization of the molecular simulation and docking techniques, we have derived the most reasonable orientations and binding positions, from which irrational possibilities were prior to that excluded. Furthermore, with the substitution study with shaker residues, the more precise roles of this area in gating have been analyzed.
Superposition of the structures of drk1 S3C and Hanatoxin before and after docking revealed a significant movement of S3C in the direction presumably towards S4. This was then comprehended as a possible factor to interfere S4 translocation during the gating occurs. From our observation, it is also suggested that shaker S3C does not show a Hanatoxin binding strong enough to induce the conformational change that was considered able to interfere S4 translocation. Thus, the structural change of S3C should play a very crucial role in the inhibition event, which may probably disagree with an assumption that a longer S3-S4 linker can compensate such effects, especially when the range of S4-displacement is taken into consideration. A possible molecular mechanism to illustrate this is therefore proposed.
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