Expression in Escherichia coli: production and purification of both subunits of the human general transcription factor TFIIE.
Both subunits of the human class II transcription factor TFIIE (rTFIIE alpha and rTFIIE beta) have been overexpressed in Escherichia coli at 26 degrees C using a T7 RNA polymerase expression system and further purified to apparent homogeneity. As in this system rTFIIE alpha was poorly expressed and copurified with a truncated form, we expressed rTFIIE alpha as a fusion protein. These overexpressed subunits of TFIIE are similar to the endogenous subunits according to the following criteria: molecular weight, microsequencing, and transcription activity.
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Laboratoire de Génétique Moléculaire des Eucaryotes du CNRS, U184 de Biologie Moléculaire et de Génie Génétique de l'INSERM, Faculté de Médecine, Strasbourg, France.
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Egly
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D000345:Affinity Labels; D000595:Amino Acid Sequence; D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases; D004926:Escherichia coli / Q000235:genetics; D006639:Histidine; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D019473:Transcription Factors, TFII; D014764:Viral Proteins
D000345:Affinity Labels; D010455:Peptides; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D019473:Transcription Factors, TFII; D014764:Viral Proteins; C066644:transcription factor TFIIE; C033223:polyhistidine; D006639:Histidine; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases