title: High expression of water-soluble recombinant antigenic domains of Toxoplasma gondii secretory organelles.
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.
authors.affiliation: Department of Parasitology, College of Medicine, The Catholic University of Korea, Seoul 137-701, Korea.
authors.forename: Zhaoshou
authors.initials: Z
authors.lastname: Yang
authors.affiliation: Department of Parasitology, College of Medicine, The Catholic University of Korea, Seoul 137-701, Korea.
authors.forename: Hye-Jin
authors.initials: HJ
authors.lastname: Ahn
authors.affiliation: Department of Parasitology, College of Medicine, The Catholic University of Korea, Seoul 137-701, Korea.
authors.forename: Ho-Woo
authors.initials: HW
authors.lastname: Nam
mesh_terms: D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology; D000915:Antibody Affinity; D000953:Antigens, Protozoan / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D015870:Gene Expression; D007074:Immunoglobulin G / Q000097:blood / Q000276:immunology; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D012698:Serologic Tests / Q000379:methods; D012995:Solubility; D014122:Toxoplasma / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014123:Toxoplasmosis / Q000175:diagnosis
chemical_list: D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D007074:Immunoglobulin G; D011994:Recombinant Proteins
keywords: GRA2; GRA3; IgG avidity; MIC2; ROP2; Toxoplasma gondii; antigenic domain; bioinformatics; intrinsically unstructured domain