A novel cyclic GMP-dependent protein kinase is expressed in the ring stage of the Plasmodium falciparum life cycle.
Cyclic GMP-dependent protein kinases (PKGs) are the major mediators of the cGMP signal transduction pathway and regulate a variety of physiological effects. We report here the characterization of an unusual PKG from the human malaria parasite Plasmodium falciparum (designated PfPKG). The 97.5 kDa protein contains some of the structural features of mammalian PKGs but, uniquely, contains a third predicted cGMP binding site and a degenerate fourth. Using both protein kinase activity assays and Western blotting with native P. falciparum proteins, we demonstrate here that PfPKG is expressed predominantly in the ring stage of the life cycle, suggesting a role in the development of asexual blood stage parasites. An Escherichia coli-derived recombinant protein (PfPKG2, Met115-Phe853) was purified and shown to have phosphotransferase activity in terms of both substrate phosphorylation and auto-phosphorylation. This activity was stimulated at least fivefold by 1.0 microM cyclic GMP, but was not stimulated by cAMP or by 8-pCPT-cGMP, which is a potent activator of mammalian PKGs. Several protein kinase inhibitors exhibited a range of inhibitory effects on PfPKG activity. Biochemical analysis therefore shows that PfPKG is distinct from mammalian PKGs with respect to both cyclic nucleotide analogue activation and inhibition profiles.
Deng
Wensheng
W
Department of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT, UK.
Baker
David A
DA
D000595:Amino Acid Sequence; D000818:Animals; D017869:Cyclic GMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D017125:Genes, Protozoan; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D009712:Nucleotides, Cyclic / Q000737:chemistry / Q000378:metabolism; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000254:growth & development / Q000502:physiology; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
D009712:Nucleotides, Cyclic; D011994:Recombinant Proteins; D017869:Cyclic GMP-Dependent Protein Kinases