Mutations in the aspartate receptor of Escherichia coli which affect aspartate binding.

The effects of five mutations at arginines 64, 69, and 73 of the Tar protein were analyzed using swarm assays, aspartate binding in purified membranes, and methylation both in vitro and in vivo. The defects in the responses of these mutant receptors to aspartate were shown to be directly attributable to reduced binding of aspartate to the receptor rather than to defects in their signaling characteristics. Mutations at residues 64, 69, and 73 reduced aspartate binding by factors of greater than 10(-4), 10(-3), and 10(-2), respectively. Once aspartate was bound, the mutants exhibited normal signaling properties. No cooperativity was observed in the coupling of aspartate binding to methylation, indicating that the monomers of the receptor dimer act independently. The in vitro methylation system was thus shown to be an effective way of measuring aspartate binding constants and examining the functional integrity of the proteins. The maltose responses of the receptor proteins were affected slightly, or not at all, in an in vivo methylation assay. Two models for the roles of these arginine residues in receptor function are discussed.