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Characterization of sequence-specific DNA binding by the transcription factor Oct-1
T Lundbäck1, J F Chang, K Phillips
1Department of Biochemistry and Molecular Biology, University College London, UK.
Biochemistry
|June 20, 2000
Summary
The Oct-1 POU protein undergoes partial folding when binding DNA, causing a larger heat change than expected. This binding involves six charged contacts with the DNA phosphate backbone.
Area of Science:
- Molecular Biology
- Biochemistry
- Structural Biology
Background:
- The Oct-1 transcription factor's POU domain binds DNA's octamer element, regulating gene expression.
- This domain's two parts encircle DNA to recognize the eight-base pair sequence.
Purpose of the Study:
- To characterize the binding thermodynamics of the Oct-1 POU domain to an octamer element using isothermal titration calorimetry.
- To investigate the reasons behind the observed heat capacity change during complex formation.
Main Methods:
- Isothermal titration calorimetry (ITC) was employed to measure binding thermodynamics.
- Proteolysis studies were conducted on the free and DNA-bound Oct-1 POU protein.
Main Results:
- Oct-1 POU/DNA complex formation exhibited a large negative heat capacity change (ΔCp, obs), exceeding predictions based on buried surface area.
- Ionic strength dependence indicated six charged contacts between the protein and DNA phosphate groups.
- Proteolysis data supported partial protein folding upon DNA binding.
Conclusions:
- The significant heat capacity change is primarily attributed to the partial folding of the Oct-1 POU protein during DNA complex formation.
- The findings align with structural data of Oct-1 POU/DNA complexes, confirming specific protein-DNA interactions.