A Designed Zn ²⁺ Sensor Domain Transmits Binding Information to Transmembrane Histidine Kinases

Generating stimulus-responsive allosteric signaling <i>de novo</i> is a significant challenge in protein design. In natural systems like bacterial histidine kinases (HKs), signal transduction occurs when ligand binding initiates a signal that is amplified across biological membranes over long distances to induce large-scale rearrangements and phosphorylation relays. Here, we ask whether our understanding of protein design and multidomain, intramolecular signaling has progressed sufficiently to enable engineering of a HK with tunable <i>de novo</i> components. We generated <i>de novo</i> metal-binding sensor domains and substituted them for the native sensor domain of a transmembrane HK, affording chimeras that transduce signals initiated from a <i>de novo</i> sensor. Signaling depended on the designed sensor's stability and the interdomain linker's phase and length. These results show the usefulness of <i>de novo</i> design to elucidate the biochemical mechanisms and principles of transmembrane signaling.