A continuous fluorescence assay for cyclic nucleotide phosphodiesterase hydrolysis of cyclic GMP.
The fluorescent 2'-methylanthraniloyl derivative of cyclic GMP undergoes a 45% decrease in fluorescence when it is cleaved by brain phosphodiesterase in the presence of calmodulin. This fluorescence decrease is dependent upon calcium, calmodulin, and phosphodiesterase, and correlates well (r = 0.996) with the disappearance of substrate as monitored by high-performance liquid chromatography. The Kd values determined by this fluorescence method and HPLC suggest that cyclic GMP and its fluorescent derivative exhibit similar kinetic parameters in their hydrolysis.
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000201:enzymology; D002147:Calmodulin / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000032:analysis; D005456:Fluorescent Dyes; D006868:Hydrolysis; D007700:Kinetics; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D062367:ortho-Aminobenzoates / Q000032:analysis
D002147:Calmodulin; D005456:Fluorescent Dyes; D062367:ortho-Aminobenzoates; C052862:2'-methylanthraniloyl guanosine cyclic monophosphate; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D006152:Cyclic GMP