Nucleic Acids Research, 1992, Vol. 20, No. 13 3391-3396
© 1992
MOLECULAR BIOLOGY |
A comparison of the DNA bending activities of the DNA binding proteins CRP and TFIID
Eukaryotic Gene Organisation and Expression Laboratory, Imperial Cancer Research Fund PO Box 123, Lincoln's Inn Fields, London WC2A 3PX, UK 1School of Biochemistry, University of Birmingham PO Box 363, Birmingham B15 2TT, UK
Received March 27, 1992. Revised June 11, 1992. Accepted June 11, 1992.
Protein-induced DNA bending is of importance in the formation of complex nucleoprotein assemblies such as those involved in the initiation of DNA replication or transcription initiation. We have compared the DNA bending characteristics of the Escherichla coli cyclic AMP receptor protein (CRP or CAP), an archetypal DNA bending protein, to those of TFIID, the eukaryotic TATA-element binding transcription factor. By altering the helical phasing between a CRP binding site and the E.coli melR promoter we have mapped a DNA sequence-directed bend in the downstream region of the promoter. This intrinsic DNA bend may be important in the regulation of the melR promoter by CRP in vivo. Gel retardation assays and DNAse I footprinting show that human TFIID binds to the melR promoter -10 region. Taking advantage of this fact, and using the CRP-induced DNA bend as a standard, we have employed phase sensitive detection to show that the DNA bend angle induced by TFIID is far less than that induced by CRP. Further evidence to support this conclusion comes from a comparison of the relative mobilities of CRP-DNA and TFIID-DNA complexes. These results place limits on the role of any DNA bending induced by TFIID alone in the initiation of transcription.
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