Abstract
In this paper we report the development of a recombinant strain of the yeast Pichia pastoris, which secretes an anti-carcinoembryonic antigen single chain Fv (scFv) antibody fragment to the culture supernatant as a biologically active protein, at levels of 1.2 g l(-1). The yeast scFv was purified by IMAC, with a final yield of approximately 0.440 g of 93% pure scFv per liter of culture supernatant. The specific activity in ELISA of the yeast scFv was almost three times higher than that of a bacterial periplasmic counterpart. These results reaffirm that the yeast P. pastoris is a suitable host for high level production of scFv antibody fragments with potential in vivo diagnostic and therapeutic applications.
MeSH terms
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Adenosine Triphosphatases*
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Animals
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Bacterial Proteins / genetics
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Bacterial Proteins / metabolism
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Biotechnology / methods
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Carcinoembryonic Antigen / biosynthesis*
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Carcinoembryonic Antigen / genetics*
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Carcinoembryonic Antigen / isolation & purification
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Chromatography, Affinity
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DNA-Binding Proteins*
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Enzyme-Linked Immunosorbent Assay
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Escherichia coli Proteins*
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Fermentation
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Gene Expression Regulation, Fungal
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Immunoglobulin Fragments / biosynthesis*
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Immunoglobulin Fragments / genetics*
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Immunoglobulin Fragments / isolation & purification
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Immunoglobulin Variable Region / biosynthesis
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Immunoglobulin Variable Region / genetics
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Immunoglobulin Variable Region / isolation & purification
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Mice
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MutS DNA Mismatch-Binding Protein
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Pichia / genetics
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Pichia / metabolism*
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Protein Engineering / methods
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Recombinant Proteins / biosynthesis*
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Recombinant Proteins / genetics
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Recombinant Proteins / isolation & purification
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Transformation, Genetic
Substances
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Bacterial Proteins
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Carcinoembryonic Antigen
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DNA-Binding Proteins
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Escherichia coli Proteins
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Immunoglobulin Fragments
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Immunoglobulin Variable Region
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Recombinant Proteins
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immunoglobulin Fv
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Adenosine Triphosphatases
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MutS DNA Mismatch-Binding Protein
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MutS protein, E coli