NUCLEIC ACID FOOTPRINTING OF INTERACTION OF TATA BINDING PROTEIN (TBP) WITH DNA

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TBP is a component of the nucleo-protein complexes required for the initiation of transcription by each of the three eukaryotic RNA polymerases. The binding of the TATA Binding Protein (TBP) to specific promoter sequences called TATA boxes is a key step in the initiation of transcription of genes transcribed by RNA polymerase II. The nucleotide se-quence of TATA boxes is variable among naturally occurring promoters. The application of synchrotron x-ray nucleic acid and protein footprinting to protein-DNA interactions is being addressed through the binding of the TATA Binding Protein (TBP) to specific promoter se-quences. This protein is unusual in that it induces a dramatic and drastic conformational change in the structure of the DNA; of particular interest is the introduction of two kinks in Hthe DNA by the intercalation of two pairs of phenylalanine residues. HIt has been proposed based on other types of kinetics data, that these Hintercalation events may proceed sequentially and that their rates may Hbe DNA sequence dependent. Titration experiments have been Hsuc-cessfully conducted which demonstrate that accurate synchrotron Hfootprinting data can be acquired for this system. The stage is now Hset for the initiation of synchrotron footprinting ki-netics Hexperiments that will critically test this hypothesis.