Luciferase-streptavidin fusion proteins: Preparation and properties

We constructed plasmids encoding genes of fusion proteins of a thermostable mutant of Luciola mingrelica firefly luciferase (Luc) and streptavidin (SA) with the polyhistidine sequence (His₆) at the N- or C-terminus of the protein: SA-Luc-His₆, His₆-SA-Luc, Luc-SA-His₆. Fusion proteins were produced and purified; their composition, luciferase activity, thermal stability, bioluminescence spectra, and biotin-binding capacity were investigated. Streptavidin introduction does not affect the bioluminescence spectra, but it decreases the thermal stability twofold at 47°C. It was shown by size-exclusion chromatography that, depending on the plasmid structure, the fusion proteins were expressed as dimers, tetramers, and larger oligomers, which differ in luciferase activity and biotin-binding capacity. The His₆-SA-Luc fusion protein demonstrated the most optimal properties.