title: Evidence of the presence of a calmodulin-sensitive plasma membrane Ca2+-ATPase in Trypanosoma equiperdum.
abstract: Trypanosoma equiperdum belongs to the subgenus Trypanozoon, which has a significant socio-economic impact by limiting animal protein productivity worldwide. Proteins involved in the intracellular Ca2+regulation are prospective chemotherapeutic targets since several drugs used in experimental treatment against trypanosomatids exert their action through the disruption of the parasite intracellular Ca2+homeostasis. Therefore, the plasma membrane Ca2+-ATPase (PMCA) is considered as a potential drug target. This is the first study revealing the presence of a PMCA in T. equiperdum (TePMCA) showing that it is calmodulin (CaM) sensitive, revealed by ATPase activity, western-blot analysis and immuno-absorption assays. The cloning sequence for TePMCA encodes a 1080 amino acid protein which contains domains conserved in all PMCAs so far studied. Molecular modeling predicted that the protein has 10 transmembrane and three cytoplasmic loops which include the ATP-binding site, the phosphorylation domain and Ca2+translocation site. Like all PMCAs reported in other trypanosomatids, TePMCA lacks a classic CaM binding domain. Nevertheless, this enzyme presents in the C-terminal tail a region of 28 amino acids (TeC28), which most likely adopts a helical conformation within a 1-18 CaM binding motif. Molecular docking between Trypanosoma cruzi CaM (TcCaM) and TeC28 shows a significant similarity with the CaM-C28PMCA4b reference structure (2kne). TcCaM-TeC28 shows an anti-parallel interaction, the peptide wrapped by CaM and the anchor buried in the hydrophobic pocket, structural characteristic described for similar complexes. Our results allows to conclude that T. equiperdum possess a CaM-sensitive PMCA, which presents a non-canonical CaM binding domain that host a 1-18 motif.
authors.affiliation: Instituto de Biología Experimental (IBE), Universidad Central de Venezuela (UCV), Caracas, Venezuela.
authors.forename: María Carolina
authors.initials: MC
authors.lastname: Pérez-Gordones
authors.affiliation: Centro de Estudios Biomédicos y Veterinarios, Instituto de Estudios Científicos y Tecnológicos (IDECYT), Universidad Nacional Experimental Simón Rodríguez (UNESR), Caracas, Venezuela.
authors.forename: José Rubén
authors.initials: JR
authors.lastname: Ramírez-Iglesias
authors.affiliation: Instituto de Biología Experimental (IBE), Universidad Central de Venezuela (UCV), Caracas, Venezuela.
authors.forename: Vincenza
authors.initials: V
authors.lastname: Cervino
authors.affiliation: Facultad de Ciencias Naturales y Ambientales, Universidad Internacional SEK (UISEK), Quito, Ecuador.
authors.forename: Graciela L
authors.initials: GL
authors.lastname: Uzcanga
authors.affiliation: Instituto de Biología Experimental (IBE), Universidad Central de Venezuela (UCV), Caracas, Venezuela; Instituto de Estudios Avanzados (IDEA), Caracas, Venezuela.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Benaim
authors.affiliation: Centro de Estudios Biomédicos y Veterinarios, Instituto de Estudios Científicos y Tecnológicos (IDECYT), Universidad Nacional Experimental Simón Rodríguez (UNESR), Caracas, Venezuela. Electronic address: mendozamarta17@gmail.com.
authors.forename: Marta
authors.initials: M
authors.lastname: Mendoza
mesh_terms: D000595:Amino Acid Sequence; D015153:Blotting, Western; D000252:Calcium-Transporting ATPases / Q000032:analysis* / Q000737:chemistry / Q000235:genetics; D002147:Calmodulin / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology*; D003001:Cloning, Molecular; D007118:Immunoassay; D008958:Models, Molecular; D011446:Prospective Studies; D011487:Protein Conformation; D000072417:Protein Domains; D016415:Sequence Alignment; D014345:Trypanosoma / Q000201:enzymology* / Q000235:genetics
chemical_list: D002147:Calmodulin; D000252:Calcium-Transporting ATPases
keywords: Animal trypanosomiasis; Ca(2+)-ATPase; Calmodulin; PMCA; Trypanosoma equiperdum