Characterization of the chemoreceptors in the Bacillus subtilis chemotaxis system.

摘要

The two-component chemotaxis signal transduction system provides peritrichously flagellated bacteria with the ability to sense their surrounding chemical environment and move towards more favorable conditions. At the core of this system are transmembrane chemoreceptors that sense these environmental cues and transmit this signal to the histidine kinase CheA. Once activated, CheA interacts with the response regulator CheY, which causes a change in the rotation of the flagella. Cells can bias their motion in chemical gradients by altering their duration of smooth swims and tumbles, governed by the rotation of the flagella, so that they can move up a favorable chemical gradient. Adaptation to these conditions is achieved in the B. subtilis chemotaxis system by three systems: the methylation system, the CheC/CheD/CheY-P system and the CheV system. Coordination of these processes is vital to the chemotactic ability of the bacteria.;The sensing domain of the chemoreceptors is responsible for binding an attractant ligand. In E. coli, the structure of this domain has been characterized as a four-helix bundle. The B. subtilis sensing domain, using structural modeling, was found to be vastly different. It contains two PAS domain, and represents a novel architecture for chemoreceptor sensing. There are specific residues responsible for ligand binding in the upper PAS domain, and mutation of any one of these residues led to a defect in the chemotactic response and a significantly lower affinity for attractant ligand.;Post-translational modification of the chemoreceptors is necessary for both chemotactic ability and adaptation. The function of the CheD deamidase was shown to be critical to the ability of B. subtilis to perform chemotaxis. Its deamidase activity was decoupled from its role in the CheC/CheD/CheY-P adaptational system, and it appeared that both roles are necessary for proper chemotactic behavior. It was also shown that deamidation of the receptor can lead to a more active chemoreceptor. Furthermore, the effect of the methylation system was analyzed and shown to be necessary for chemotactic function.;Physical characterization of the chemoreceptors was probed by identification of a zinc ion binding area, using isothermal titration calorimetry. In addition, in vivo chemotactic assays showed that altering the charges of the methylation sites on the chemoreceptors can have a significant effect on the activity of the receptor. Furthermore, individual methylation sites were shown to be responsible for both changes in the affinity of the chemoreceptor for attractant and receptor activation. These findings suggest a novel, more complicated role for methylation than previously suggested, and lead to a clearer picture of the coordination of the three adaptational systems in the B. subtilis chemotactic sensory transduction.