Korean J Parasitol.  2014 Aug;52(4):367-376. 10.3347/kjp.2014.52.4.367.

High Expression of Water-Soluble Recombinant Antigenic Domains of Toxoplasma gondii Secretory Organelles

• Yang Z
• Ahn HJ
• Nam HW

• KMID: 1792197
• DOI: http://doi.org/10.3347/kjp.2014.52.4.367

Abstract

Recombinant antigenic proteins of Toxoplasma gondii are alternative source of antigens which are easily obtainable for serodiagnosis of toxoplasmosis. In this study, highly antigenic secretory organellar proteins, dense granular GRA2 and GRA3, rhoptrial ROP2, and micronemal MIC2, were analyzed by bioinformatics approach to express as water-soluble forms of antigenic domains. The transmembrane region and disorder tendency of 4 secretory proteins were predicted to clone the genes into pGEX-4T-1 vector. Recombinant plasmids were transformed into BL21 (DE3) pLysS E. coli, and GST fusion proteins were expressed with IPTG. As a result, GST fusion proteins with GRA225-105, GRA339-138, ROP2324-561, and MIC21-284 domains had respectively higher value of IgG avidity. The rGST-GRA225-105 and rGST-GRA339-138 were soluble, while rGST-ROP2324-561 and rGST-MIC21-284 were not. GRA231-71, intrinsically unstructured domain (IUD) of GRA2, was used as a linker to enhance the solubility. The rGST-GRA231-71-ROP2324-561, a chimeric protein, appeared to be soluble. Moreover, rGST-GRA231-71-MIC21-284 was also soluble and had higher IgG avidity comparing to rGST-MIC21-284. These 4 highly expressed and water-soluble recombinant antigenic proteins may be promising candidates to improve the serodiagnosis of toxoplasmosis in addition to the major surface antigen of SAG1.

Keyword

Toxoplasma gondii; bioinformatics; intrinsically unstructured domain; GRA2; GRA3; ROP2; MIC2; antigenic domain; IgG avidity

MeSH Terms

Animals
Antibodies, Protozoan/immunology
Antibody Affinity
Antigens, Protozoan/chemistry/*diagnostic use/genetics/immunology
*Gene Expression
Immunoglobulin G/blood/immunology
Mice, Inbred BALB C
Recombinant Proteins/chemistry/*diagnostic use/genetics/immunology
Serologic Tests/methods
Solubility
Toxoplasma/genetics/immunology/*metabolism
Toxoplasmosis/diagnosis
Antibodies, Protozoan
Antigens, Protozoan
Immunoglobulin G
Recombinant Proteins