BRET-SENSORMOLEKÜLE ZUR DETEKTION VON HYDROLASEN

摘要

The present invention relates to bioluminescence resonance energy transfer sensor molecules having the structure R1-L-R2 -B or B- R2-L-R1, wherein R1 is a bioluminescent protein, L is a linking element, R2 is a non-protein acceptor domain and B is a blocking group, and wherein R2 bound to B comprises a hydrolysable bond which produces a change in BRET when hydrolysed. The invention also discloses a method of detecting a hydrolase by contacting a sample with a molecule B-R2, then contacting with a compound R1-L or L-R1 under conditions to cause attaching of R2 to L, and detecting a change in the BRET ratio. Specifically exemplified sensors comprise luciferase and fluorescein diacetate, which is hydrolysed by an esterase. The invention also discloses luciferase enzymes derived from RLuc8 by removing cysteine residues.