Nonspecific vs. specific DNA binding free energetics of a transcription factor domain protein

Transcription factor (TF) proteins regulate gene expression by binding to specific sites on the genome. In the facilitated diffusion model, an optimized search process is achieved by the TF alternating between 3D diffusion in the bulk and 1D diffusion along DNA. While undergoing 1D diffusion, the protein can switch from a search mode for fast diffusion along nonspecific DNA to a recognition mode for stable binding to specific DNA. It was recently noticed that, for a small TF domain protein, reorientations on DNA happen between the nonspecific and specific DNA binding. We here conducted all-atom molecular dynamics simulations with steering forces to reveal the protein-DNA binding free energetics, confirming that the search and recognition modes are distinguished primarily by protein orientations on the DNA. As the binding free energy difference between the specific and nonspecific DNA system slightly deviates from that being estimated directly from dissociation constants on 15-bp DNA constructs, we hypothesize that the discrepancy can come from DNA sequences flanking the 6-bp central binding sites that impact on the dissociation kinetics measurements. The hypothesis is supported by a simplified spherical protein-DNA model along with stochastic simulations and kinetic modeling.