PREPARATION OF A GENETICALLY FUSED PROTEIN-A LUCIFERASE CONJUGATE FOR USE IN BIOLUMINESCENT IMMUNOASSAYS

The genes encoding staphylococcal protein A and bacterial luciferase (Vibrio harveyi) were fused in-frame in order to obtain a general marker enzyme for bioluminescent immunoassays. Two constructs were made where protein A was ligated to the first and the 12th amino acid residue, respectively, of the N terminus of the beta-subunit of luciferase. Only the first fusion protein encoding the entire beta-subunit was able to form an enzymatically active luciferase complex when expressed together with the alpha-subunit. The fusion of protein A to luciferase did not notably alter the emitted wavelength spectrum or its stability to urea treatment. The fusion protein was found to retain at least 50% of the specific bioluminescent activity compared to native luciferase. In preliminary tests, this hybrid protein was shown to be useful in bioluminescent immunoassays.