FRET protease assays for clostridial toxins
AssigneeAllergan, Inc.
InventorsLance E. Steward, Ester Fernandez-Salas, Kei Roger Aoki
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Abstract
The present invention provides clostridial toxin substrates useful in assaying for the protease activity of any clostridial toxin, including botulinum toxins of all serotypes as well as tetanus toxins. A clostridial toxin substrate of the invention contains a donor fluorophore; an acceptor having an absorbance spectrum overlapping the emission spectrum of the donor fluorophore; and a clostridial toxin recognition sequence that includes a cleavage site, where the cleavage site intervenes between the donor fluorophore and the acceptor and where, under the appropriate conditions, resonance energy transfer is exhibited between the donor fluorophore and the acceptor.
Claims15
A botulinum toxin substrate, comprising:
- a) a donor fluorophore;
- b) an acceptor having an absorbance spectrum overlapping the emission spectrum of said donor fluorophore, wherein the acceptor is an acceptor fluorophore;
- and
- c) a botulinum toxin type B recognition sequence comprising a botulinum toxin type B P5-P4-P3-P2-P1-P1′-P2′-P3′-P4′-P5′ cleavage site sequence, said botulinum toxin type B P5-P4-P3-P2-P1-P1′-P2′-P3′-P4′-P5′ cleavage site sequence intervening between said donor fluorophore and said acceptor;
- wherein either of said donor fluorophore, said acceptor, or both said donor fluorophore and said acceptor are genetically encoded;
- and
- wherein, under the appropriate conditions, resonance energy transfer is exhibited between said donor fluorophore and said acceptor fluorophore.
Dependencies are inferred from the claim text.