A potent voltage-gated calcium channel inhibitor engineered from a nanobody targeted to auxiliary CaVβ subunits

• DOI: 10.7554/elife.49253
• Publication Date: 2019-08-12T12:00:19Z
• Authors: Travis J Morgenstern; Jinseo Park; Qing R Fan; Henry M Colecraft
• Keywords: HEK 293 cells; Transport protein

Inhibiting high-voltage-activated calcium channels (HVACCs; CaV1/CaV2) is therapeutic for myriad cardiovascular and neurological diseases. For particular applications, genetically-encoded HVACC blockers may enable channel inhibition with greater tissue-specificity versatility than achievable small molecules. Here, we engineered a inhibitor by first isolating an immunized llama nanobody (nb.F3) that binds auxiliary CaVβ subunits. Nb.F3 itself functionally inert, providing convenient vehicle to target active moieties CaVβ-associated channels. fused the catalytic HECT domain of Nedd4L (CaV-aβlator), E3 ubiquitin ligase, ablated currents from diverse HVACCs reconstituted in HEK293 cells, endogenous CaV1/CaV2 mammalian cardiomyocytes, dorsal root ganglion neurons, pancreatic β cells. In CaV-aβlator redistributed CaV1.2 dyads Rab-7-positive late endosomes. This work introduces as potent inhibitor, describes general approach can be broadly adapted generate versatile modulators macro-molecular membrane protein complexes.