ENZYME ASSAY FOR MUTANT FIREFLY LUCIFERASE

摘要

Enzymes and methods suitable for assaying ATP, and specific applications for such assays are described and claimed. In particular, there is described a recombinant mutant luciferase having a mutation (e.g. the amino-acid corresponding to amino acid residue number 245 in Photinus pyralis which is such that the Km for ATP of the luciferase is increased e.g. five-fold with respect to that of the corresponding non-mutated enzyme such that it is of the order of 500 .mu.m - 1mM. Also disclosed are luciferases having additional mutations conferring improved thermostability or altered wavelength of emitted light. Recombinant polynucloetides, vectors and host cells are also disclosed, as are methods of assaying the amount of ATP in a material (e.g. cells) optionally in real-time. Also disclosed are test-kits for in vitro assays.