Characterization and binding analysis of a microneme adhesive repeat domain-containing protein from Toxoplasma

Parasitol Int. 2013 Dec 18. pii: S1383-5769(13)00207-9. doi: 10.1016/j.parint.2013.12.006. [Epub ahead of print]

Characterization and binding analysis of a microneme adhesive repeat domain-containing protein from Toxoplasma gondii

Gong H1, Kobayashi K2, Sugi T3, Takemae H4, Ishiwa A3, Recuenco FC3, Murakoshi F2, Xuan X4, Horimoto T2, Akashi H2, Kato K5.

Author information

Abstract

The intracellular parasite Toxoplasma gondii invades almost all nucleated cells, and has infected approximately 34% of the world's population to date. In order to develop effective vaccines against T. gondii infection, understanding of the role of the molecules that are involved in the invasion process is important. For this purpose, we characterized T. gondii proteins that contain microneme adhesive repeats (MAR), which are common in moving junction proteins. T. gondii MAR domain-containing protein 4a (TgMCP4a), which contains repeats of 17-22 amino acid segments at the N-terminus and three putative MAR domains at the C-terminus, is localized near the rhoptry of extracellular parasites. Following infection, TgMCP4a was detected in the parasitophorous vacuole. The recombinant Fc-TgMCP4a N-terminus protein (rTgMCP4a-1/Fc) showed binding activity to the surface proteins of Vero, 293T, and CHO cells. The recombinant GST-TgMCP4a N-terminus protein (rTgMCP4a-1/GST), which exhibited binding activity, was used to pull down the interacting factors from 293T cell lysate, and subsequent mass spectrometry analysis revealed that three types of heat shock proteins (HSPs) interacted with TgMCP4a. Transfection of a FLAG fusion protein of TgMCP4a-1 (rTgMCP4a-1/FLAG) into 293T cell and the following immunoprecipitation with anti-FLAG antibody confirmed the interactions of HSC70 with TgMCP4a. The addition of rTgMCP4a-1/GST into the culture medium significantly affected the growth of the parasite. This study hints that T. gondii may employ HSP proteins of host cell to facilitate their growth.

Copyright © 2013. Published by Elsevier Ireland Ltd.

KEYWORDS:

Heat shock protein, TgMCP4a, Toxoplasma

PMID:

 

24361285

 

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