Artículo
High level of expression of the Toxoplasma gondii-recombinant Rop2 protein in Escherichia coli as a soluble form for optimal use in diagnosis
Registro en:
1073-6085
10.1385/MB:18:3:269
Autor
Nigro, Mónica G
Martín, Valentina
Kaufer, Federico
Carral, Liliana
Angel, Sergio O.
Pszenny, Viviana
Resumen
Fil: Nigro, Mónica. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología. Departamento de Parasitología Sanitaria; Argentina. Fil: Martin, Valentina. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología. Departamento de Parasitología Sanitaria; Argentina. Fil: Kaufer, Federico. Hospital Alemán. Laboratorio de Toxoplasmosis; Argentina. Fil: Carral, Liliana. Hospital Alemán. Laboratorio de Toxoplasmosis; Argentina. Fil: Angel, Sergio O. Instituto Nacional de Parasitología. Departamento de Parasitología Sanitaria; Argentina. Fil: Pszenny, Viviana. Instituto Nacional de Parasitología. Departamento de Parasitología Sanitaria; Argentina. The rhoptry 2 protein (Rop2) is an interesting protein of Toxoplasma gondii that is involved in the parasite invasion of host cell, it has three T-cell epitopes and high antigenic value. However, the expression of Rop2 as a recombinant protein in Escherichia coli is not an easy task, showing low levels of expression or degradation and solubility problems. Using a recombinant Rop2(196-561) fused to 6 histidine residues, we showed high levels of expression in bacteria growing in terrific broth. rRop2(196-561) was purified mainly as a soluble product and in high concentrations (approx 1 mg/mL) under native conditions (40 mM imidazol in phosphate buffer). However, after a cycle of freezing-thawing rRop2(196-561) became insoluble. When glycerol was added to 26%, immediately after purification, the protein stayed soluble after cycles of freezing-thawing. Finally, it was demonstrated that under these conditions soluble rRop2(196-561) keeps its diagnostic value in contrast with the insoluble protein.