ADP is produced by firefly luciferase but its synthesis is independent of the light emitting properties

During experiments aimed at understanding the time course of appearance of reaction products in the Photinus pyralis luciferase system, an expected co mpound with a typical nucleotide UV spectrum was isolated. According to cap illary electrophoresis (CE) analysis and H-1, P-31-NMR spectra, it was unam biguously found to be ADP, either with extracted or recombinant enzymes. Th e ADP synthesis was demonstrated by standard UV spectrophotometric methods and CE analysis. Also, the luciferase produced AMP and ATP from ADP. This r eaction was completely inhibited by Ap(5)A at 250 nM and was independent of the light emitting properties of the enzyme. The only catalytic mechanism to explain the production of ADP is an intrinsic adenylate kinase activity of luciferase. The K-m values of the AK activity are 0.3, 0.7, 0.06 mM for AMP, ADP, and ATP respectively. The multiple enzyme activities of luciferas e may be partly responsible for the complex kinetics of light emission by c hanging the nucleotide concentrations. (C) 2001 Societe francaise de biochi mie et biologic moleculaire / Editions scientifiques et medicales Elsevier SAS. All rights reserved.