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Updated: Aug 9, 2026
High-throughput Functional Screening using a Homemade Dual-glow Luciferase Assay
Published on: June 1, 2014
Firefly luciferase produces hydrogen peroxide as a coproduct in dehydroluciferyl adenylate formation
Hugo Fraga1, Diogo Fernandes, Jiri Novotny
1Centro de Investigação em Química, Departamento de Química, Faculdade de Ciências, Universidade do Porto, R. Campo Alegre 687, 4169-007 Porto, Portugal.
Abstract:
Firefly luciferase catalyzes the synthesis of H2O2 from the same substrates as the bioluminescence reaction: ATP and luciferin (D-LH2). About 80% of the enzyme-bound intermediate D-luciferyl adenylate (D-LH2-AMP) is oxidized into oxyluciferin, and a photon is emitted during this reaction. The enzyme pathway responsible for the generation of H2O2 is a side reaction in which D-LH2-AMP is oxidized into dehydroluciferyl adenylate (L-AMP). Like the bioluminescence reaction, the luciferase-catalyzed synthesis of H2O2 and L-AMP is a stereospecific process, involving only the natural D enantiomer. However, the intramolecular electron transfer postulated as essential to the light emission process is not involved in this side reaction.
