title: Molecular characterization and localization of Plasmodium falciparum choline kinase.
abstract: Generation of phosphocholine by choline kinase is important for phosphatidylcholine biosynthesis via Kennedy pathway and phosphatidylcholine biosynthesis is essential for intraerythrocytic growth of malaria parasite. A putative gene (Gene ID PF14_0020) in chromosome 14, having highest sequence homology with choline kinase, has been identified by BLAST searches from P. falciparum genome sequence database. This gene has been PCR amplified, cloned, over-expressed and characterized. Choline kinase activity of the recombinant protein (PfCK) was validated as it catalyzed the formation of phosphocholine from choline in presence of ATP. The K(m) values for choline and ATP are found to be 145+/-20 microM and 2.5+/-0.3 mM, respectively. PfCK can phosphorylate choline efficiently but not ethanolamine. Southern blotting indicates that PfCK is a single copy gene and it is a cytosolic protein as evidenced by Western immunoblotting and confocal microscopy. A model structure of PfCK was constructed based on the crystal structure of choline kinase of C. elegans to search the structural homology. Consistent with the homology modeling predictions, CD analysis indicates that the alpha and beta content of PfCK are 33% and 14%, respectively. Since choline kinase plays a vital role for growth and multiplication of P. falciparum during intraerythrocytic stages, we can suggest that this well characterized PfCK may be exploited in the screening of new choline kinase inhibitors to evaluate their antimalarial activity.
authors.affiliation: Division of Drug Target Discovery and Development, Central Drug Research Institute, Chatter Manzil Palace, Mahatma Gandhi Marg, Lucknow-226001, Uttar Pradesh, India.
authors.forename: Vinay
authors.initials: V
authors.lastname: Choubey
authors.forename: Mithu
authors.initials: M
authors.lastname: Guha
authors.forename: Pallab
authors.initials: P
authors.lastname: Maity
authors.forename: Sanjay
authors.initials: S
authors.lastname: Kumar
authors.forename: Resmi
authors.initials: R
authors.lastname: Raghunandan
authors.forename: Prakas R
authors.initials: PR
authors.lastname: Maulik
authors.forename: Kalyan
authors.initials: K
authors.lastname: Mitra
authors.forename: Umesh C
authors.initials: UC
authors.lastname: Halder
authors.forename: Uday
authors.initials: U
authors.lastname: Bandyopadhyay
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D017173:Caenorhabditis elegans; D002797:Choline Kinase / Q000737:chemistry*; D004912:Erythrocytes / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010767:Phosphorylcholine / Q000737:chemistry; D010963:Plasmodium falciparum / Q000201:enzymology*; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid
chemical_list: D010767:Phosphorylcholine; D000255:Adenosine Triphosphate; D002797:Choline Kinase