In vivo preparation of [32P]cAMP and thin-layer chromatography of cAMP-related compounds.
A simple and rapid method for preparing [32P]adenosine 3'5'-cyclic monophosphate (cAMP) is described. A culture of an Escherichia coli mutant which excretes cAMP about 150 times faster than does a wild-type strain was incubated overnight with [32P]orthophosphate of high specific activity (e.g., 4000 Ci/mol (1 Ci = 37 GBq). The [32P]cAMP which accumulated extracellularly was then purified to 99.9% radiochemical purity in less than 4 h by adsorption to charcoal and alumina column chromatography. A two-dimensional chromatography system using a PEI-cellulose plate is also described which should prove useful for studying cAMP metabolism with 32P- or 3H-labeled cAMP or ATP.
D002855:Chromatography, Thin Layer; D000242:Cyclic AMP / Q000096:biosynthesis*; D004926:Escherichia coli / Q000378:metabolism*; D007553:Isotope Labeling / Q000379:methods*; D010761:Phosphorus Radioisotopes
D010761:Phosphorus Radioisotopes; D000242:Cyclic AMP