FACTORS INVOLVED IN SPECIFIC TRANSCRIPTION BY MAMMALIAN RNA POLYMERASE-II - PURIFICATION AND CHARACTERIZATION OF GENERAL TRANSCRIPTION FACTOR TFIIE

Human transcription factor TFIIE, a ubiquitous factor required for transcription initiation by RNA polymerase II, was purified to homogeneity by a combination of conventional and HPLC steps. The purified TFIIE contained equimolar amounts of 57-kDa (TFIIE-α) and 34-kDa (TFIIE-β) polypeptides that were judged to be functional subunits on the basis of their copurification with transcriptional activity and the recovery of activity following renaturation of polypeptides separated by reverse-phase HPLC. TFIIE-α had an independent TFIIE activity whereas TFIIE-β had no activity alone but enhanced the activity of TFIIE-α. In conjunction with gel filtration studies, which indicated a molecular mass of ≈180 kDa for the native protein, these results suggested that TFIIE is a heterotetramer containing two a and two β polypeptides. Functional studies with the purified TFIIE demonstrated that it is a general initiation factor, required for all of the genes tested, but it failed to show any DNA-dependent ATPase activity.