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Abstract

Transmembrane receptors allow cells to monitor their environment and initiate appropriate responses. The molecular mechanism of signal transduction is not yet fully understood, but the successful construction of functional chimeras shows that an entire class of receptors, comprising histidine kinases, chemoreceptors, adenylate cyclases and phosphatases, shares the same mechanism. One of the smallest known receptors to date, Af1503 from Archaeoglobus fulgidus, has proven particularly suitable for modular design and

structural studies. Its HAMP domain was the first to be solved at high resolution and led to a model for signal transduction based on axial helix rotation. However, further progress was stalled by our inability to solve the full-length structure of the protein, until AlphaFold2 provided us with a sufficiently good model for molecular replacement. In contrast to other receptors from the same class, Af1503 lacks a C-terminal effector domain, making it difficult to identify its natural ligand and establish its functional role. To address this problem, we engineered a functional chimera, consisting of Af1503 fused to the EnvZ histidine kinase

domain. This chimera is well folded and fully functional in vivo, enabling us to determine its substrate specificity. In conjunction with crystallography studies, we could establish that the native ligands of Af1503 are fatty acids of 12 to 18-carbon atom chains, opening the way to study the mechanism of signal transduction at atomic resolution.