A basic feature of living cells is the ability to respond to specific, external chemical signals. Like eukaryotic cells that respond to hormones or neurotransmitters, bacteria contain specific receptors, exposed on the cell surface, that recognize relevant compounds. These cells are chemotactic as the result of the functioning of a multicomponent, sensory-response system that links receptors to flagella. The long-term goal of this laboratory is to contribute to a detailed molecular biological description of the chemotactic system. Recent progress makes it likely that this will be the first receptor system to be understood in detail. The growing number of analogies between bacterial chemoreception and eukaryotic receptor systems suggest that at least some of the information obtained about the prokaryotic system will have general significance. This proposal involves physiological, genetic and biochemical approaches to the study of chemoreception in Escherichia coli. Primary emphasis is placed on the study of transducers, which are integral membrane proteins central to both the excitation and adaptation phases of chemotactic behavior. Adaptation involves covalent modification of transducers, specifically protein carboxyl methylation at multiple glutamyl residue sites. Some of those glutamyl residues are present because glutamines are enzymatically deamidated to create glutamates. The functional significance of multiple methylation and deamidation will be examined by alteration of the modification sites using synthetic oligonucleotide-directed mutagenesis. The pattern of transducer oligomers will be studied with the hope of defining the significance of these complexes in tactic function. The structure and conformational changes of transducers will be investigated by purifying the proteins, by using spectroscopic methods to monitor conformational differences and by developing conditions in which the molecules can be crystallized.
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