Firefly luciferyl: adenylate

DeLuca, M., Dempsey, M. E., Hori, K., and Cormier, M. J. (1976). Source of oxygen in CO2 produced during chemiluminescence of firefly luciferyl-adenylate and Renilla luciferin. Biochem. Biophys. Res. Commun. 69 262-267. 

The following schemes represent the overall reaction of firefly bioluminescence (McElroy and DeLuca, 1978), where E is luciferase LH2 is D-luciferin PP is pyrophosphate AMP is adenosine phosphate LH2-AMP is D-luciferyl adenylate (an anhydride formed between the carboxyl group of luciferin and the phosphate group of AMP) and L is oxyluciferin. 

In the postulated bioluminescence mechanism, firefly luciferin is adenylated in the presence of luciferase, ATP and Mg2+. Luciferyl adenylate in the active site of luciferase is quickly oxygenated at its tertiary carbon (position 4), forming a hydroperoxide intermediate (A). 

The firefly enzyme (EC 1.13.12.7) catalyzes the intermediate formation of D(-)-luciferyl adenylate and pyrophosphate fromD(-)-luciferin and ATP, followed by the oxidative reaction of the acyl adenylate with molecular oxygen to form an enzyme-bound product in the excited... 

The structure of firefly luciferin has been confirmed by total synthesis. The firefly emits a ycllow-grccn luminescence, and luciferin in this case is a benzthiazole derivative. Activation of the firefly luciferin involves the elimination of pyrophosphate from ATP widi the formation of an add anhydride linkage between the carboxyl group of luciferin and the phosphate group of adenylic acid forming luciferyl-adenylate. 

IMPORTANCE OF FIREFLY LUCIFERASE C-TERMINAL DOMAIN IN BINDING OF LUCIFERYL-ADENYLATE... 

Firefly Luciferase C-Terminal Domain in Luciferyl-Adenylate Binding... 

Dukhovich A, Sillero A, Sillero MA. Time course of luciferyl adenylate synthesis in the firefly luciferase reaction. FEBS Lett 1996 395 188-90. 

Fraga H, Esteves da Silva JCG Fontes R. Identification of luciferyl adenylate and luciferyl coenzyme A synthesized by firefly luciferase. ChemBioChem 2004 5 110-5. 

Some years ago I demonstrated that such a biochemical flash can be produced in the test tube.9,15 If oxygen is excluded from a firefly luciferase reaction mixture and then added rapidly back, a bright flash occurs, some 100 to 200 times brighter than the baseline intensity (Fig. 3). This comes from the reaction of the luciferyl adenylate active intermediate accumulated in the absence of oxygen. Note that the decay of the flash is not due to the removal of oxygen, but to the utilization of the luciferase-peroxide intermediate, so the baseline returns to a low level (Fig. 4), defined by the slow rate of reaction of ATP with luciferin. It is well known that the kinetics of firefly flashes are species specific and of functional importance in courtship communication, fixed by the rate constant for the first order decay of the peroxide intermediate formed from the adenylate. 

The first step in the formation of light in the firefly is a reaction with ATP to form luciferyl adenylate (Eq. 23-46, step The proton on the carbon may then be removed making use of the electron accepting properties of the adjacent ring system and carbonyl group before addition of the O2. The reactions should be compared to those catalyzed by oxygenases, e.g.,... 

The luciferase from the North American firefly P. pyralis is a monomeric enzyme (62 kDa) consisting of 550 amino acid residues. The firefly luciferase produces light by the ATP-dependent oxidation of D(-)-luciferin (LH2) (see Scheme 8.3). The reaction involves an enzyme-hound luciferyl adenylate intermediate. The peak light emission occurs at 562 nm (yellow—green, quantum yield of 0.88) in dianionic form between pH 7 and 8 or at 610 nm (red, quantum yield of 0.2) in monoanionic form at pH values below 7. The red shift also occurs in the presence of Zn (2.3 mM) or Cd (12 mM) (58). 

As far as is known, all other fireflies use the same compound in essentially the same way. A notable feature is the requirement for ATP, the result being the formation of luciferyl adenylate (1). This reaction predisposes the luciferin to attack by base and oxygen, and provides the leaving group which results in the formation of the dioxetanone (2). 

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