RCK (regulating the conductance of K+) domains are ubiquitous among a wide variety of cation translocation systems or channels. Whether it is inward or outward transport, RCK domains share high sequence similarity from protein to protein, suggesting that they perform important functions in these systems. Although specific functions are not yet fully understood, RCK domains bind nucleotides, a characteristic that has been suggested to be important for the open/closed transition. In some cases RCK domains can bind additional ligands, e.g. KefC binding to glutathione. This project provides an in-depth study of two equally important bacterial potassium uptake systems, Trk (from E. coli) and CglK (from C. glutamicum). In these systems, RCK domains form octamers that either are anchored (CglK) or are separate and bind to transmembrane partners (Trk). The overall objective of this study was to examine the ligand control of the Trk potassium uptake system, including ligand identification and binding effects on its conformation, and therefore activity control. A crystal structure of the Trk potassium uptake system from Vibrio parahaemolyticus, was published recently [1] and provided evidence of ADP/ATP switch as a control mechanism. In accordance with the aforementioned study, the work presented in this thesis provides strong evidence that both ADP and ATP can bind to E. coli TrkA. Furthermore, it was shown that NADH bind strongly to one of the two RCK domains present in TrkA protein. The data presented here suggest a more complex control mechanism of the E. coli Trk system. CglK is a major potassium uptake system of C. glutamicum, but little is known about its control mechanism. The mutagenesis approach was undertaken to learn more about the system and its underlying processes/mechanisms. The work shown in this thesis indicates a similarity of CglK to other potassium channels, such as MthK and GsuK. The functional mechanisms proposed for those two systems were proven to be similar to that of CglK, although the specific CglK activation ligands are still to be found.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:606422 |
| Date | January 2013 |
| Creators | Pasko, Jaroslaw Piotr |
| Publisher | University of Aberdeen |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=210109 |
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