Abstract
The method leading to overexpression of the full-length mouse recombinant prion protein (mrPrP 23-231) in the cytoplasm of E. coli as a his-PrP fusion protein and its effective purification using affinity chromatography is described. A typical yield of the method was 8-10 mg his-mrPrP per L of the bacterial culture. The purity of purified protein was > 95 %. The purified his-mrPrP was converted to a soluble form and its folding to alpha-helical and beta-sheet conformations was studied. The properties of differently folded mrPrP were determined by measuring their circular dichroism spectra, partial resistance to cleavage by proteinase K and by centrifugation in sucrose gradient.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Blotting, Western
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Chromatography, Affinity
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Circular Dichroism
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Electrophoresis, Polyacrylamide Gel
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Escherichia coli / genetics
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Escherichia coli / metabolism
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Mice
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Oxidation-Reduction
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Peptide Fragments / biosynthesis
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Peptide Fragments / chemistry
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Peptide Fragments / genetics
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Peptide Fragments / isolation & purification*
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Prions / biosynthesis
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Prions / chemistry
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Prions / genetics
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Prions / isolation & purification*
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Protein Folding
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Recombinant Proteins / biosynthesis
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Recombinant Proteins / chemistry
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Recombinant Proteins / genetics
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Recombinant Proteins / isolation & purification
Substances
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Peptide Fragments
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Prions
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Recombinant Proteins
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prion protein (23-231)