Optimization of the expression of surface antigen SAG1/2 of Toxoplasma gondii in the yeast Pichia pastoris

Thiruvengadam, G.; Init, I.; Fong, M.Y.; Lau, Y.L. (2011) Optimization of the expression of surface antigen SAG1/2 of Toxoplasma gondii in the yeast Pichia pastoris. Tropical Biomedicine, 28 (3). pp. 506-513. ISSN 0127-5720

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    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 30 masculineC, 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.

    Item Type: Article
    Creators:
    1. Thiruvengadam, G.
    2. Init, I.(Department of Parasitology, Faculty of Medicine Building, University of Malaya, 50603 Kuala Lumpur, MALAYSIA)
    3. Fong, M.Y.
    4. Lau, Y.L.
    Journal or Publication Title: Tropical Biomedicine
    Additional Information: Thiruvengadam, G Init, I Fong, M Y Lau, Y L eng Evaluation Studies Research Support, Non-U.S. Gov't Malaysia 2012/03/22 06:00 Trop Biomed. 2011 Dec;28(3):506-13.
    Uncontrolled Keywords: Ammonium Sulfate/chemistry Antigens, Protozoan/*biosynthesis/diagnostic use/genetics/isolation & purification Carbon/metabolism Chromatography, Affinity Cloning, Molecular Culture Media/chemistry Dialysis Fractional Precipitation *Gene Expression Hydrogen-Ion Concentration Methanol/metabolism Pichia/*genetics Protozoan Proteins/*biosynthesis/diagnostic use/genetics/isolation & purification Recombinant Fusion Proteins/biosynthesis/diagnostic use/genetics/isolation & purification Sensitivity and Specificity Serologic Tests/methods Temperature Toxoplasmosis/diagnosis
    Subjects: R Medicine
    Divisions: Faculty of Medicine
    Depositing User: Mr Jenal S
    Date Deposited: 06 Dec 2012 14:57
    Last Modified: 06 Dec 2012 14:57
    URI: http://eprints.um.edu.my/id/eprint/4076

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