title: Optimization of the expression of surface antigen SAG1/2 of Toxoplasma gondii in the yeast Pichia pastoris.
abstract: Surface antigens are the most abundant proteins found on the surface of the   parasite Toxoplasma gondii. Surface antigen 1 (SAG1) and Surface antigen 2 (SAG2) remain   the most important and extensively studied surface proteins. These antigens have been   identified to play a role in host cell invasion, immune modulation, virulence attenuation.   Recombinant SAG1/2 was cloned and expressed in yeast Pichia pastoris. We describe here   optimization of critical parameters involved in high yield expression of the recombinant   SAG1/2. Our results suggest that recombinant SAG1/2 were best expressed at 30ºC, pH 6 and   1% methanol as the carbon source by X33 Pichia cells. Additional optimizations included the   downstream process such as ammonium sulphate precipitation and dialysis. The fusion protein   was purified using Ni-NTA purification system with 80% recovery. The purified protein was   100% specific and sensitive in detection of toxoplasmosis.
authors.affiliation: School of Science, Monash University Sunway Campus, Bandar Sunway, 46150 Selangor, Malaysia.
authors.forename: G
authors.initials: G
authors.lastname: Thiruvengadam
authors.forename: I
authors.initials: I
authors.lastname: Init
authors.forename: M Y
authors.initials: MY
authors.lastname: Fong
authors.forename: Y L
authors.initials: YL
authors.lastname: Lau
mesh_terms: D000645:Ammonium Sulfate / Q000737:chemistry; D000953:Antigens, Protozoan / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D002244:Carbon / Q000378:metabolism; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003470:Culture Media / Q000737:chemistry; D003956:Dialysis; D005590:Fractional Precipitation; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D000432:Methanol / Q000378:metabolism; D010843:Pichia / Q000235:genetics*; D015800:Protozoan Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D012698:Serologic Tests / Q000379:methods; D013696:Temperature; D014123:Toxoplasmosis / Q000175:diagnosis
chemical_list: D000953:Antigens, Protozoan; D003470:Culture Media; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; C047168:SAG1 antigen, Toxoplasma; C067547:surface antigen P22, Toxoplasma; D002244:Carbon; D000645:Ammonium Sulfate; D000432:Methanol