Membrane Association Transforms an Inert Anti-TCRβ Fab' Ligand into a Potent T Cell Receptor Agonist

The natural peptide-major histocompatibility complex (pMHC) ligand for T cell receptors (TCRs) is inactive from solution yet capable of activating T cells at single-molecule levels when membrane-associated. This distinctive feature stems from the mechanism of TCR activation, which is thought to involve steric phosphatase exclusion as well as direct mechanical forces. It is possible to defeat this mechanism and activate T cells with solution ligands by cross-linking pMHC or using multivalent antibodies to TCR. However, these widely used strategies activate TCRs through a nonphysiological mechanism and can produce different activation profiles than natural, monovalent, membrane-associated pMHC. Here, we introduce a strictly monovalent anti-TCRβ H57 Fab' ligand that, when coupled to a supported lipid bilayer via DNA complementation, triggers TCRs and activates nuclear translocation of the transcription factor nuclear factor of activated T cells (NFAT) with a similar potency to pMHC in primary murine T cells. Importantly, like monovalent pMHC and unlike bivalent antibodies, monovalent Fab'-DNA triggers TCRs only when physically coupled to the membrane, and only around 100 individual Fab':TCR interactions are necessary to stimulate early T cell activation.