Abstract

Neurological and endocrine pathologies such as acromegalie, Cushing’s disease, and
neuropathic pain display disregulated exocytosis. Silencing specific cell populations would
thus be invaluable to correct these debilitating disorders. To achieve this goal, we reengineered
the Botulinum neurotoxin (BoT), a highly potent pharmaceutical compound
capable of inhibiting exocytosis, and fused to it a protein “stapling” domain [1,2]. These
peptide motifs, that form an irreversible tetrahelical coiled-coil, are able to link a variety of
targeting domains onto the enzyme and thus redirect it towards normally unaffected cells.
The conformational diversity of this assembly process greatly supersedes traditional protein
expression since multiple targeting domains (homo- and hetero-) can be linked onto one
scaffold, larger yields can be produced separately, it permits the combination of solid-phase
peptide synthesis with recombinant protein expression, and it can avoid the necessity of an
N- to C- translational fusion. With only a few dozen building “blocks” it is possible to
construct thousands of different complexes specifically tailored for each purpose as every
individual component can be linked onto any other cognate stapling moieties.