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.
Jepson
S G
SG
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