The process by which bacteria bias their motility, enabling them to move towards favourable chemical stimuli and away from unfavourable ones, is called chemotaxis. Motility and chemotaxis are essential for the virulence of many medically important bacteria, including Helicobacter pylori, and also play a role in agriculturally important bacteria, such as the nitrogen-fixing Azospirillum brasilense. The chemotaxis signalling network of E. coli has been extensively studied. The small 14kD protein CheY is an important response regulator in this chemotaxis signalling network, and conserved across all systems. It undergoes a conformational change when activated by phosphorylation which increases its affinity for the FliM component of the flagellar motor switch complex. This results in a change in the direction of the flagellar motor rotation. Most bacteria have much more complex chemosensory systems than those of E. coli. Rhodobacter sphaeroides, the subject of this proposal, has multiple homologues of the E. coli chemosensory proteins. For example, it produces six homologues of the response regulator CheY. These CheY’s, CheY1/2/3/4/5/6, are localized to and are regulated by different clusters of chemosensory proteins in the cell and, while they all bind the motor switch, they have different effects on chemotaxis. The aim of this project is to understand, at the level of individual amino acids, how these six highly homologous CheY proteins are fine-tuned to carry out their specific roles. State-of-the-art NMR methods will be used to study the structure and dynamics of several CheY’s in their inactive and active states. Low populations of ‘active-like’ conformations that exist in unphosphorylated, inactive CheY’s will be characterized using NMR relaxation dispersion methods. These ‘invisible’ states are likely to play a crucial role in the activation mechanism of the CheY’s, and differences observed for the six homologues will help in understanding their specific roles.
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