LIGAND & SUBUNIT INTERACTIONS OF TRANSMEMBRANE RECEPTORS

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DESCRIPTION: Studies are proposed of the aspartate and serine receptor proteins from the Escherichia coli chemotaxis pathway, and of the cytoplasmic signalling proteins with which they interact, regarding issues of linkage, cooperativity, conformational change and clustering using quantitative thermodynamic (isothermal titration calorimetry) and spectroscopic methods (Fluorescence and EPR) with biochemical assays of signalling activity. A feature of many transmembrane receptors is the dimeric, or more generally oligomeric, organization of binding permits the system to exhibit cooperativity and linkage phenomena that have important consequences for many membrane phenomena. Under physiological conditions it is not understood the extent to which transmembrane signalling occurs by changes in receptor clustering, through conformational changes within stable clusters of receptor, or both. These issues could be resolved if more information were available about the conformational changes, the clustering reactions, and the mutual influence of ligands binding on opposite sides of the membrane (transbilayer binding effects). It is proposed that this information will be obtained through (i) precise measurements of the thermodynamic and kinetic parameters that characterize ligand binding, domain interaction and subunit interaction of membrane proteins, (ii) accurate and sensitive spectroscopic methods to determine changes in receptor conformation and dynamics in response to ligand binding and protein-protein interactions, and (iii) localization of the sites of interaction between signalling proteins. Completion of the proposed studies will give a complete picture of the physical mechanism of transmembrane signalling and the molecular mechanism of receptor desensitization by covalent modification. The bacterial chemotaxis pathway belongs to the superfamily of two-component signalling systems that have been found in prokaryotes, eukaryotic microorganisms and plants. The information should thus be of widespread fundamental significance, and also directly relevant to understanding signal transduction processes in pathogenic microbes.