Characterization of EhCaBP, a calcium-binding protein of Entamoeba histolytica and its binding proteins.
A novel calcium-binding protein (EhCaBP) has been recently identified and characterized from the protozoan parasite Entamoeba histolytica. In order to decipher the function of this protein, a few basic properties were investigated and compared with the ubiquitous Ca(2+)-signal transducing protein calmodulin (CaM). Indirect immunofluorescence and immunoprecipitation analyses using specific antibodies against EhCaBP suggest that it is a soluble cytoplasmic protein with no major post-translational modification. EhCaBP did not stimulate cAMP-phosphodiesterase activity, differentiating it from all known CaMs. Affinity chromatography of [35S]methionine-labelled proteins of E. histolytica trophozoites using EhCaBP-sepharose column showed Ca(2+)-dependent binding of a group of proteins. Radiolabelled proteins from the same extract also bound to CaM-sepharose. However, the proteins bound to the two columns were different as revealed by sodium dodecyl sulphate polyacrylamide gel electrophoresis. At least one of the EhCaBP-binding proteins became phosphorylated as revealed by in vivo phosphorylation analysis. The binding-proteins could not be detected in E. invadens (a species that is pathogenic in reptiles) and E. moshkovskii (which is found in the human gut but is not pathogenic), two species in which EhCaBP-like protein has not been found. Two distinct Ca(2+)-dependent protein kinases, which get activated by EhCaBP and CaM respectively, were detected in E. histolytica. These kinases require different levels of Ca2+ for their maximal activities. Affinity chromatography also showed the binding of protein kinase(s) to EhCaBP in a Ca(2+)-dependent manner. Our data suggest that there may be novel Ca(2+)-signal transduction pathway in E. histolytica mediated by EhCaBP.
Yadava
N
N
School of Life Sciences, Jawaharlal Nehru University, New Delhi, India.
Chandok
M R
MR
Prasad
J
J
Bhattacharya
S
S
Sopory
S K
SK
Bhattacharya
A
A
D000818:Animals; D002135:Calcium-Binding Proteins / Q000302:isolation & purification* / Q000378:metabolism; D004748:Entamoeba histolytica / Q000737:chemistry*; D004789:Enzyme Activation; D011494:Protein Kinases / Q000032:analysis* / Q000378:metabolism; D015800:Protozoan Proteins / Q000032:analysis* / Q000302:isolation & purification* / Q000378:metabolism
D002135:Calcium-Binding Proteins; D015800:Protozoan Proteins; C075554:calcium-binding protein, Entamoeba histolytica; D011494:Protein Kinases