title: Characterization of Plasmodium falciparum calcium-dependent protein kinase 1 (PfCDPK1) and its role in microneme secretion during erythrocyte invasion.
abstract: Calcium-dependent protein kinases (CDPKs) play important roles in the life cycle of Plasmodium falciparum and other apicomplexan parasites. CDPKs commonly have an N-terminal kinase domain (KD) and a C-terminal calmodulin-like domain (CamLD) with calcium-binding EF hands. The KD and CamLD are separated by a junction domain (JD). Previous studies on Plasmodium and Toxoplasma CDPKs suggest a role for the JD and CamLD in the regulation of kinase activity. Here, we provide direct evidence for the binding of the CamLD with the P3 region (Leu(356) to Thr(370)) of the JD in the presence of calcium (Ca(2+)). Moreover, site-directed mutagenesis of conserved hydrophobic residues in the JD (F363A/I364A, L356A, and F350A) abrogates functional activity of PfCDPK1, demonstrating the importance of these residues in PfCDPK1 function. Modeling studies suggest that these residues play a role in interaction of the CamLD with the JD. The P3 peptide, which specifically inhibits the functional activity of PfCDPK1, blocks microneme discharge and erythrocyte invasion by P. falciparum merozoites. Purfalcamine, a previously identified specific inhibitor of PfCDPK1, also inhibits microneme discharge and erythrocyte invasion, confirming a role for PfCDPK1 in this process. These studies validate PfCDPK1 as a target for drug development and demonstrate that interfering with its mechanistic regulation may provide a novel approach to design-specific PfCDPK1 inhibitors that limit blood stage parasite growth and clear malaria parasite infections.
authors.affiliation: International Centre for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi 110067, India.
authors.forename: Abhisheka
authors.initials: A
authors.lastname: Bansal
authors.forename: Shailja
authors.initials: S
authors.lastname: Singh
authors.forename: Kunal R
authors.initials: KR
authors.lastname: More
authors.forename: Dhiraj
authors.initials: D
authors.lastname: Hans
authors.forename: Kuldeep
authors.initials: K
authors.lastname: Nangalia
authors.forename: Manickam
authors.initials: M
authors.lastname: Yogavel
authors.forename: Amit
authors.initials: A
authors.lastname: Sharma
authors.forename: Chetan E
authors.initials: CE
authors.lastname: Chitnis
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000595:Amino Acid Sequence; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000235:genetics / Q000378:metabolism; D003514:Cyclohexylamines / Q000494:pharmacology; D004912:Erythrocytes / Q000187:drug effects / Q000469:parasitology; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D052940:Merozoites / Q000187:drug effects / Q000201:enzymology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015388:Organelles / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology; D010963:Plasmodium falciparum / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D002147:Calmodulin; D003514:Cyclohexylamines; D010455:Peptides; D047428:Protein Kinase Inhibitors; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C531239:purfalcamine; D011494:Protein Kinases; C531238:calcium-dependent protein kinase-1, Plasmodium falciparum; D000225:Adenine; D002118:Calcium