Enhancement of firefly luciferase activity by cytidine nucleotides.
The temporal pattern of light production by firefly luciferase depends on the ATP concentration. With low concentrations of ATP a constant production of light occurred while at high concentrations of ATP (greater than 10 microM) there was a flash of light followed by a decline in light production. This time course of light production with high ATP concentrations was changed from the flash pattern to a pattern with a constant production of light by several cytidine nucleotides. CTP, CDP, dCTP, dCDP, dideoxyCTP, periodate-oxidized CTP and CDP, and the etheno derivatives of CTP and CDP produced that change. CMP, cytidine, CDP-glycerol, CDP-glucose, CDP-ethanolamine, and benzoylbenzoylCTP either were inhibitory to firefly luciferase or were not effective in changing the flash time course. Coenzyme A and related compounds also changed the time course of light production. The changes in time course produced by either cytidine nucleotides or CoA were inhibited by desulfoCoA. These compounds apparently enhanced light production by promoting the dissociation of the inhibitory product, oxidized luciferin, from the enzyme. When the activating compounds were used with high concentrations of ATP, the sensitivity of assay for firefly luciferase was increased. This increased sensitivity is important when using the firefly luciferase gene as a reporter.
Ford
S R
SR
Department of Biochemistry, Oklahoma State University, Stillwater 74078-0454.
Hall
M S
MS
Leach
F R
FR
D000227:Adenine Nucleotides / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D003065:Coenzyme A / Q000737:chemistry / Q000378:metabolism; D003597:Cytosine Nucleotides / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008156:Luciferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009987:Orthoptera / Q000201:enzymology; D010084:Oxidation-Reduction; D010504:Periodic Acid / Q000737:chemistry; D013329:Structure-Activity Relationship
D000227:Adenine Nucleotides; D003597:Cytosine Nucleotides; D010504:Periodic Acid; D000255:Adenosine Triphosphate; D008156:Luciferases; D003065:Coenzyme A