Presence of a thapsigargin-sensitive calcium pump in Trypanosoma evansi: Immunological, physiological, molecular and structural evidences.
Exp Parasitol
; 159: 107-17, 2015 Dec.
Article
en En
| MEDLINE
| ID: mdl-26297682
ABSTRACT
In higher eukaryotes, the sarco-endoplasmic reticulum (ER) Ca(2+)-ATPase (SERCA) is characterized for its high sensitivity to low concentrations of thapsigargin (TG), a very specific inhibitor. In contrast, SERCA-like enzymes with different sensitivities to TG have been reported in trypanosomatids. Here, we characterized a SERCA-like enzyme from Trypanosoma evansi and evaluated its interaction with TG. Confocal fluorescence microscopy using BODIPY FL TG and specific anti-SERCA antibodies localized the T. evansi SERCA-like enzyme in the ER and confirmed its direct interaction with TG. Moreover, the use of either 1 µM TG or 25 µM 2',5'-di (tert-butyl)-1,4-benzohydroquinone prevented the reuptake of Ca(2+) and consequently produced a small increase in the parasite cytosolic calcium concentration in a calcium-free medium, which was released from the ER pool. A 3035 bp-sequence coding for a protein with an estimated molecular mass of 110.2 kDa was cloned from T. evansi. The corresponding gene product contained all the invariant residues and conserved motifs found in other P-type ATPases but lacked the calmodulin binding site. Modeling of the three-dimensional structure of the parasite enzyme revealed that the amino acid changes found in the TG-SERCA binding pocket do not compromise the interaction between the enzyme and the inhibitor. Therefore, we concluded that T. evansi possesses a SERCA-like protein that is inhibited by TG.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Trypanosoma
/
ATPasas Transportadoras de Calcio
/
Bombas Iónicas
/
Tapsigargina
/
Inhibidores Enzimáticos
Tipo de estudio:
Diagnostic_studies
Límite:
Animals
Idioma:
En
Revista:
Exp Parasitol
Año:
2015
Tipo del documento:
Article
País de afiliación:
Venezuela