Firefly luciferase is able to convert (L)-luciferin into luciferyl-CoA even under ordinary aerobic luciferin-luciferase reaction conditions. The luciferase is able to recognize strictly the chirality of the luciferin structure, serving as the acyl-CoA synthetase for (L)-luciferin, whereas (D)-luciferin is used for the bioluminescence reaction. (D)-Luciferin inhibits the luciferyl-CoA synthetase activity Of (L)-luciferin, whereas (L)-luciferin retards the bioluminescence reaction Of (D)-luciferin, meaning that both enzyme activities are prevented by the enantiomer of its own substrate. © 2005 Elsevier Inc. All rights reserved.