Abstract
Motile Escherichia coli cells use chemoreceptor signaling arrays to track chemical gradients with exquisite precision. Highly conserved residues in the cytoplasmic hairpin tip of chemoreceptor molecules promote assembly of trimer-based signaling complexes and modulate the activity of their CheA kinase partners. To explore hairpin tip output states in the serine receptor Tsr, we characterized the signaling consequences of amino acid replacements at the salt-bridge residue pair E385-R388. All mutant receptors assembled trimers and signaling complexes, but most failed to support serine chemotaxis in soft agar assays. Small side-chain replacements at either residue produced OFF- or ON-shifted outputs that responded to serine stimuli in wild-type fashion, suggesting that these receptors, like the wild-type, operate as two-state signaling devices. Larger aliphatic or aromatic side chains caused slow or partial kinase control responses that proved dependent on the connections between core signaling units that promote array cooperativity. In a mutant lacking one of two key adapter-kinase contacts (interface 2), those mutant receptors exhibited more wild-type behaviors. Lastly, mutant receptors with charged amino acid replacements assembled signaling complexes that were locked in kinase-ON (E385K|R) or kinase-OFF (R388D|E) output. The hairpin tips of mutant receptors with these more aberrant signaling properties probably have nonnative structures or dynamic behaviors. Our results suggest that chemoeffector stimuli and adaptational modifications influence the cooperative connections between core signaling units. This array remodeling process may involve activity-dependent changes in the relative strengths of interface 1 and 2 interactions between the CheW and CheA.P5 components of receptor core signaling complexes.