Plant Physiol. Drug Metab Dispos
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Plant Physiology 65:165-170 (1980)
© 1980 American Society of Plant Biologists

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Articles

Synthesis and Release of Cyclic Adenosine 3':5'-Monophosphate by Ochromonas malhamensis1

Ray A. Bressan2, Avtar K. Handa2, Hartmut Quader3 and Philip Filner

MSU-DOE Plant Research Laboratory, Michigan State University, East Lansing, Michigan 48824

The chrysophycean alga, Ochromonas malhamensis Pringsheim, was shown to synthesize cyclic adenosine 3':5'-monophosphate (cAMP) and to release it into the culture medium. Cells contained 3 to 3,000 picomoles per gram fresh weight; medium contained up to 20 times the amount in the cells. Putative [32P]cAMP was purified from cultures supplied [32P]phosphate. The compound was identified as [32P]cAMP by co-chromatography with authentic cAMP through 10 serial steps; by chemical deamination at the same rate as authentic cAMP, to a 32P compound with the chromatographic behavior of cIMP; and by its conversion through the action of cyclic nucleotide phosphodiesterase to a 32P compound with the chromatographic behavior of 5'-AMP. A two-step procedure involving chromatography on alumina and on Dowex 50 purified the unlabeled compound from cells or medium sufficiently for it to be assayable by competitive inhibition of binding of [3H]cAMP to cAMP-binding protein (Gilman assay) or by stimulation of cAMP-dependent protein kinase. The activity was destroyed by cyclic nucleotide phosphodiesterase with the same kinetics as authentic cAMP, provided that an endogenous inhibitor of the phosphodiesterase was first removed by an additional purification step.


2 Present address: Department of Horticulture, Purdue University, Lafayette, Indiana 47907.

3 Present address: Pflanzenphysiologisches Institut der Universität, Abtlg Cytologie, Untere Karlspüle 2, D-3400 Göttingen, West Germany. Supported by a fellowship from the DFG.

1 This work was supported by United States Department of Energy Contract EY-76-C-02-1338.







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