Nucleotide sequence, expression, and properties of luciferase coded by lux genes from a terrestrial bacterium.
The lux genes required for expression of luminescence have been cloned from a terrestrial bacterium, Xenorhabdus luminescens, and the nucleotide sequences of the luxA and luxB genes coding for the alpha and beta subunits of luciferase determined. The lux gene organization was closely related to that of marine bacteria from the Vibrio genus with the luxD gene being located immediately upstream and the luxE downstream of the luciferase genes, luxAB. A high degree of homology (85% identity) was found between the amino acid sequences of the alpha subunits of X. luminescens luciferase and the luciferase from a marine bacterium, Vibrio harveyi, whereas the beta subunits of the two luciferases had only 60% identity in amino acid sequence. The similarity in the sequences of the alpha subunits of the two luciferases was also reflected in the substrate specificities and turnover rates with different fatty aldehydes supporting the proposal that the alpha subunit almost exclusively controls these properties. The luciferase from X. luminescens was shown to have a remarkably high thermal stability being stable at 45 degrees C (t 1/2 greater than 3 h) whereas V. harveyi luciferase was rapidly inactivated at this temperature (t 1/2 = 5 min). These results indicate that the X. luminescens lux system may be the bacterial bioluminescent system of choice for application in coupled luminescent assays and expression of lux genes in eukaryotic systems at higher temperatures.
Szittner
R
R
Department of Biochemistry, McGill University, Montreal, Quebec, Canada.
Meighen
E
E
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D004755:Enterobacteriaceae / Q000201:enzymology / Q000235:genetics*; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D008156:Luciferases / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid
D004269:DNA, Bacterial; D008156:Luciferases