Protease activity and host cell binding of the 42-kDa rhoptry protein from Toxoplasma gondii after secretion.
Three proteases were identified in the excretory/secretory proteins (ESP) from Toxoplasma gondii by the gelatin acrylamide gel electrophoresis (GAGE), of which the molecular masses were 80, 70, and 42 kDa. One of the proteases with 42 kDa was reactive to a monoclonal antibody (mAb), Tg786 clone, which was localized in the rhoptry of T. gondii by immunohistochemistry. The protease was maximally active at the pH range between 7.5 and 8.5, and was sensitive to inhibition by TPCK and EGTA. The gelatinolytic activity of the protease was dependent on the concentration of calcium ion. The protease was active only in the millimolar ranges of calcium but not in micromolar ranges, implicating that the secretion is critical event for the activation of the protease. The secreted protease was shown to bind to the host cells upon Western blot and immunofluorescence analysis. It is suggested that the protease may target to the plasma membrane of the host cells, which provides appropriate environment for the entry of the parasite into host cells. The mAb (Tg786) of T. gondii also reacted with a protein of the same size and equivalent locality of rhoptry in Neospora caninum, a similar Apicomplexan protozoa, suggesting that secreted protease mediates a common function in the mechanism of entry into host cells.
Ahn
H J
HJ
Department of Parasitology and Catholic Institute of Parasitic Diseases, Catholic University of Korea, Seoul, Korea.
Song
K J
KJ
Son
E S
ES
Shin
J C
JC
Nam
H W
HW
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007150:Immunohistochemistry; D007477:Ions; D051379:Mice; D008807:Mice, Inbred BALB C; D008856:Microscopy, Fluorescence; D011233:Precipitin Tests; D011485:Protein Binding; D013997:Time Factors; D014122:Toxoplasma / Q000201:enzymology* / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D007477:Ions; D002118:Calcium