Cyclic AMP- and Ca2(+)-dependent protein kinases in Plasmodium falciparum.
The cyclic AMP- and Ca2(+)-dependent protein kinase activities of Plasmodium falciparum were partially characterized after purification of parasites from host erythrocytes by N2 cavitation and Percoll gradient centrifugation. Proteins of molecular weights 80, 54, 51, and 31.5 kDa were phosphorylated in a cAMP-dependent manner in cytosolic extracts of isolated P. falciparum. Cytosolic extracts also contained cAMP-dependent histone II-A kinase activity with an average Vmax of 131.1 pmol/32P/min/mg protein and a Km for cAMP of 85nM. Upon photoaffinity labeling with [32P]-8-N3-cAMP, a 53-kDa protein was specifically labeled in parasite cytosol. A metabolically labeled protein of the same molecular weight was identified by cAMP-agarose affinity chromatography. The 53-kDa protein cochromatographed with cAMP-dependent histone II-A kinase activity on DEAE-cellulose, suggesting that it is the regulatory subunit of the kinase. Ca2(+)-dependent phosphorylation of proteins of molecular weights 195, 158, 51, 47.5, and 15 kDa was demonstrated in a membrane fraction from parasites free of the erythrocyte membrane. This activity was not stimulated by either calmodulin or phospholipid plus diacylglycerol and was absent from the membranes of uninfected erythrocytes. Of several exogenous substrates tested, none were found to be a substrate for this Ca2(+)-dependent kinase. Both cAMP- and Ca2(+)-dependent kinases phosphorylated serine and threonine residues.
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Department of Cellular and Molecular Physiology, Tufts University School of Medicine, Boston, Massachusetts.
Mikkelsen
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D000818:Animals; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D003600:Cytosol / Q000201:enzymology; D004912:Erythrocytes / Q000201:enzymology / Q000469:parasitology; D006801:Humans; D008970:Molecular Weight; D010766:Phosphorylation; D010963:Plasmodium falciparum / Q000201:enzymology*; D011494:Protein Kinases / Q000032:analysis* / Q000302:isolation & purification; D017871:Calcium-Calmodulin-Dependent Protein Kinases
D011494:Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases