Characterization of Temperature-Dependent Gating in Archaebacterial Calcium Activated Potassium Channel

Abstract

MthK is an archaebacterial K+ channel, which is gated by large cytoplasmic RCK domains that facilitate Ca2+-activation of the channel. This bacterial channel is a good model for studying the mechanisms underlying calcium dependent gating in ion channels because it tractable to both structural and functional studies. An earlier study has also suggested that the calcium-dependent gating of MthK is also regulated by temperature (Parfenova, L. V. et al. (2006). J Biol Chem 281, 21131-21138). Here, using inside-out patch-clamping recording of inactivation removed MthK from spheroplasts, I have characterized the temperature-dependence of MthK channel at varying Ca2+ concentrations. Our preliminary data shows that the activation of this inactivation-removed channel is highly temperature-dependent. We have also purified and reconstituted this inactivation removed MthK in soybean polar lipids and found similar temperature-dependence albeit at lower concentrations of calcium. This suggests that the temperature-dependence of MthK is intrinsic. Furthermore, allosteric analysis of electrophysiological data shows that temperature unexpectedly modulates coupling between Ca2+ sensor and the pore rather than acting through a completely independent allosteric domain.