Notch signalling is a conserved pathway that regulates cell fate decisions and is initiated by binding of a transmembrane ligand (Jagged (JAG) or Delta-like (DLL)) to a Notch receptor. This binding is followed by proteolytic cleavage of the receptor, which requires molecular tension. Building on previous studies of DLL4–Notch1 interactions, Luca et al. applied in vitro protein evolution to generate a high-affinity JAG1 variant, and, using crystallography and affinity analysis, demonstrated that different ligands interact with Notch in clearly distinct ways. DLL4 binds to Notch with higher affinity than does JAG1, and lower tension was required for the activation of Notch by DLL4 or high-affinity JAG1 than by wild-type JAG1. These results suggest that mechanical tuning of Notch signalling occurs through the modulation of ligand–receptor affinity, which is determined by their intermolecular interactions.