title: New fluorescent analogs of cAMP and cGMP available as substrates for cyclic nucleotide phosphodiesterase.
abstract: The synthesis of fluorescent derivatives of cAMP and cGMP, by reaction with isatoic anhydride in aqueous solution at mild pH and temperature, yielding 2'-O-anthraniloyl derivatives of cyclic nucleotides, is here described. 2'-O-(N-Methylanthraniloyl) derivatives were also synthesized by reaction with N-methylisatoic anhydride. Upon excitation at 330-350 nm, these derivatives exhibited maximum fluorescence emission at 430-445 nm in aqueous solution with quantum yields of 0.11-0.26. Their fluorescence was sensitive to the polarity of solvent; in N,N-dimethylformamide quantum yields of 0.8-0.95. The major differences between the two fluorophores were the longer wavelength of the emission maximum of the N-methylanthraniloyl group and its greater quantum yield. The derivatives were substrates for beef heart cyclic nucleotide phosphodiesterase, 15-24% as effective as the natural substrate cAMP. When combined with thin layer chromatography techniques, two apparent Km values (3-4 microM and 36-76 microM) for the cAMP derivatives and one value (10-18 microM) for the cGMP derivatives were obtained. The results indicate that these 2'-hydroxyl-modified cAMP and cGMP can be useful fluorescent substrate analogs for cyclic nucleotide phosphodiesterase.
mesh_terms: D015087:2',3'-Cyclic-Nucleotide Phosphodiesterases / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D000242:Cyclic AMP / Q000031:analogs & derivatives*; D006152:Cyclic GMP / Q000031:analogs & derivatives*; D005456:Fluorescent Dyes; D007700:Kinetics; D009206:Myocardium / Q000201:enzymology*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D051381:Rats; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship
chemical_list: D005456:Fluorescent Dyes; D000242:Cyclic AMP; D015087:2',3'-Cyclic-Nucleotide Phosphodiesterases; D010727:Phosphoric Diester Hydrolases; D006152:Cyclic GMP