A response regulator is paired specifically to its protein kinase. The response regulator generally has two domains. The N-terminus is generally the receiver domain and is approximately 120 amino acids in length. The conserved sequences, including three aspartyl and one lysine residue, define membership of the response regulator family. Folding of the domain brings the conserved residues into juxtaposition forming the active site. The receiver domain catalyzes transfer of the phosphoryl group from the histidine of the sensor kinase to a conserved aspartate residue. It can also utilize a variety of small molecules (but not ATP) as alternative phosphodonors. The phosphorylation state of the receiver modulates the activity of the unique C-terminal output domain, commonly a transcriptional regulator, to elicit the adaptive response. Most of the response regulators have potential helix-turn-helix DNA-binding motifs in their C-terminal domains.

The stability of the phosphorylated response regulators depends on the presence or absence of intrinsic autophosphatase activity and varies widely with half-lives ranging from 10 s for CheY to 1 hour for OmpR.

The role of phosphorylation in the activation of response regulators may be to stabilise a conformational change in the protein. Certainly, a modified CheY with Asp-13 replaced by lysine is not phosphorylated, yet phenotypically it seems to be in an active conformation, resembling the wild-type phosphorylated CheY.