Expression, Purification and Investigation of luminescence Decay in Bacterial Luciferase Xenorhabdus luminescence by Replacement of Glu175 by Gly on α Subunit

Expression, purification and comparison of fluorescence quenching in native and mutant bacterial luciferase at different concentration of FMN has been investigated. Bacterial luciferase consists of two-subunit α and β. that are encoded by the LuxA and LuxB genes, respectively. Kinetic properties of bacterial luciferase are determined by α subunit. Bacterial luciferases catalyzes the light emission reaction (Wavelength 490 nm) utilizing FMNH2, fatty aldehyde and O2 substrates. Bacterial luciferases (Lux AB) can be readily classed as slow or fast decay luciferases based on their rates of luminescence decay in a single turnover assay. Recent studies using random mutagenesis on α subunit in the “slow decay” bacterial luciferase Xenorhabdus luminescence, showed one of the mutants contains Glu175 replacement by Gly has a significantly more rapid decay rate luminescence than native luciferase. Expression, purification and kinetic studies of native and mutant luciferase indicated that difference between native and mutant luciferase in luminescence decay rate may occur of weaker binding of FMN to enzyme in mutant luciferase