A Novel Transcription Initiation Factor (TIF), TIF-IE, Is Required for Homogeneous Acanthamoeba castellanii TIF-IB (SL1) to Form a Committed Complex

• Published in: The Journal of biological chemistry Vol. 273; no. 42; pp. 27708 - 27715
• Main Authors: Radebaugh, Catherine A., Kubaska, William M., Hoffman, Laura H., Stiffler, Kristine, Paule, Marvin R.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 16-10-1998; American Society for Biochemistry and Molecular Biology
• Subjects: Acanthamoeba - genetics; Acanthamoeba castellanii; Animals; Centrifugation, Zonal; Chromatography, Affinity; DNA, Protozoan - metabolism; DNA-Binding Proteins - isolation & purification; DNA-Binding Proteins - metabolism; Freshwater; Pol1 Transcription Initiation Complex Proteins; Promoter Regions, Genetic; Protein Binding; RNA Polymerase I - metabolism; TATA-Box Binding Protein; Transcription Factors - isolation & purification; Transcription Factors - metabolism; Transcription, Genetic
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.273.42.27708

The fundamental transcription initiation factor (TIF) for ribosomal RNA expression by eukaryotic RNA polymerase I, TIF-IB, has been purified to near homogeneity fromAcanthamoeba castellanii using standard techniques. The purified factor consists of the TATA-binding protein and four TATA-binding protein-associated factors with relative molecular weights of 145,000, 99,000, 96,000, and 91,000. This yields a calculated native molecular weight of 460,000, which compares well with its mass determined by scanning transmission electron microscopy (493,000) and its sedimentation rate, which is close to RNA polymerase I (515,000). Both impure and nearly homogeneous TIF-IB exhibit an apparent equilibrium dissociation constant of 56 ± 3 pm. However, although impure TIF-IB can form a promoter-DNA complex resistant to challenge by other promoter-containing DNAs, near homogeneous TIF-IB cannot do so. An additional transcription factor, dubbed TIF-IE, restores the ability of near homogeneous TIF-IB to sequester DNA into a committed complex.