Structural gating studies of the KcsA cytoplasmic domain
We are interested in how the KcsA cytoplasmic C-terminal domain (CTD), a homotetrameric 40-residue long helical bundle, modulates the opening of the channel in response to pH changes. Contrary to the prevalent dogma, we showed using NMR and sedimentation equilibrium that CTD itself contains a pH-sensor which causes it to dissociate at lower pH levels, and that the 'pH switch' is the critical H145 residue with a pKa of ~6, rather than the acidic residues of CTD. In order to properly assess these findings in the cellular context we are now investigating this behavior in full-length channels embedded in lipoprotein nanodiscs and advanced isotopic labeling methods.
Kamnesky, G.; Shaked, H.; Chill, J.H. The distal C-terminal region of the KcsA potassium channel is a pH-dependent tetramerization domain. J. Mol. Biol. 2012, 418(3-4), 237-247.
Kamnesky, G.; Hirschhorn, O.; Shaked, H.; Chen, J.; Yao, L.; Chill, J.H. Molecular determinants of tetramerization in the KcsA cytoplasmic domain. Prot. Science, 2014, 23(10):1403-1416.
Qasim, A.; Sher I.; Hirschhorn O.; Shaked H.; Qasem Z.; Ruthstein S.; Chill J. H., Investigation of a KcsA cytoplasmic pH gate in lipoprotein nanodiscs. ChemBioChem 2019, 20, 813.