Purification of a maize dehydrin protein expressed in Escherichia coli.
A maize dehydrin protein (Dhn1) containing 167 amino acids with a predicted molecular weight of 17.0 kDa was produced in the Escherichia coli overexpression strain BL21 (DE3)pLysS. Site-directed mutagenesis was used to construct a plasmid with a protein coding region corresponding exactly to the original cDNA. Protein production was induced by IPTG. Dhn1 was enriched from total soluble protein by heat-fractionation and centrifugation and then purified by sequential cation exchange and hydrophobic interaction chromatography. The purified protein was visualized by SDS-PAGE and immunoblot analysis using a polyclonal antibody to the dehydrin consensus region. Expression in E. coli resulted in approximately 1.2 mg of purified protein per liter of induced culture.
Department of Botany and Plant Sciences, University of California, Riverside 92521-0124, USA.
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D000595:Amino Acid Sequence; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange / Q000379:methods; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015151:Immunoblotting; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification; D012995:Solubility; D013997:Time Factors; D003313:Zea mays / Q000737:chemistry*
D010940:Plant Proteins; D011994:Recombinant Proteins; C070556:dehydrin proteins, plant