| Ion channels are integral membrane proteins that selectively and efficiently catalyze the transfer of ions down an electrochemical gradient. The functions of these channels are associated with many basic cellular functions, including the regulation of electrical activity in excitable cells. This thesis focuses on understanding fundamental molecular mechanisms behind gating and permeation in potassium channels using an approach that relies on structural and functional analysis of biochemically tractable prokaryotic channels as models for their evolutionary descendants.;The first results focus on the characterization of the NaK ion channel from B. cereus. These results reveal that NaK contains a novel structural element, which confers current rectification and is an important modulator of channel gating. After this characterization, the focus shifted to a search for a physiological activator of NaK. Although none was found, results show that iron binds to NaK in a conformation-dependent manner. The final part of this thesis focuses on the engineering of the NaK channel as a structural scaffold for a human potassium channel, hERG. Results show that the lack of interactions between amino acids in the pore region of the channel gives rise to conformational flexibility, which helps to explain the nature of hERG's rapid inactivation.;The result of the work presented here is an advance in understanding the structure-function relationship in ion channels. Not only are new phenomena revealed through analysis of a previously uncharacterized channel, NaK, but also new insight is given to help explain established phenomena in a well-studied channel, hERG. Most importantly, this work sets the stage for new questions to be asked in order to advance the field of ion channel research. |
