1 research outputs found
Thermodynamic Additivity for Impacts of Base-Pair Substitutions on Association of the Egr‑1 Zinc-Finger Protein with DNA
The transcription factor Egr-1 specifically
binds as a monomer
to its 9 bp target DNA sequence, GCGÂTÂGÂGÂGCG,
via three zinc fingers and plays important roles in the brain and
cardiovascular systems. Using fluorescence-based competitive binding
assays, we systematically analyzed the impacts of all possible single-nucleotide
substitutions in the target DNA sequence and determined the change
in binding free energy for each. Then, we measured the changes in
binding free energy for sequences with multiple substitutions and
compared them with the sum of the changes in binding free energy for
each constituent single substitution. For the DNA variants with two
or three nucleotide substitutions in the target sequence, we found
excellent agreement between the measured and predicted changes in
binding free energy. Interestingly, however, we found that this thermodynamic
additivity broke down with a larger number of substitutions. For DNA
sequences with four or more substitutions, the measured changes in
binding free energy were significantly larger than predicted. On the
basis of these results, we analyzed the occurrences of high-affinity
sequences in the genome and found that the genome contains millions
of such sequences that might functionally sequester Egr-1