Affinity purification of a human RNA polymerase II complex using monoclonal antibodies against transcription factor IIF.
Five different monoclonal antibodies that immunoreact with RAP74, the large subunit of general transcription factor (TF) IIF, were produced and characterized. Using one of these antibodies, an affinity purification procedure was devised to isolate a human RNA polymerase II complex. This procedure is fast, simple, and reproducible and does not require extensive purification. The RNA polymerase II complex isolated using this procedure contains SRB (suppressor of RNA polymerase B) polypeptides, transcription factors IIE and IIF, limiting amounts of TFIIH, and the TATA-binding protein, but was devoid of TFIIB.
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Howard Hughes Medical Institute, Division of Nucleic Acid Enzymology, Department of Biochemistry, University of Medicine and Dentistry of New Jersey, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
Maldonado
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Reinberg
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D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D002846:Chromatography, Affinity / Q000379:methods*; D004268:DNA-Binding Proteins / Q000378:metabolism; D018604:Epitope Mapping; D006801:Humans; D008969:Molecular Sequence Data; D011485:Protein Binding; D012319:RNA Polymerase II / Q000302:isolation & purification* / Q000378:metabolism; D035181:TATA-Box Binding Protein; D035581:Transcription Factor TFIIB; D051758:Transcription Factor TFIIH; D014157:Transcription Factors / Q000032:analysis* / Q000276:immunology* / Q000378:metabolism; D019473:Transcription Factors, TFII; D014158:Transcription, Genetic
D000911:Antibodies, Monoclonal; D004268:DNA-Binding Proteins; D035181:TATA-Box Binding Protein; D035581:Transcription Factor TFIIB; D014157:Transcription Factors; D019473:Transcription Factors, TFII; C066644:transcription factor TFIIE; D051758:Transcription Factor TFIIH; D012319:RNA Polymerase II; C056604:transcription factor TFIIF