Complete Coupled Binding-Folding Pathway of the Intrinsically Disordered Transcription Factor Protein Brinker Revealed by Molecular Dynamics Simulations and Markov State Modeling.

Intrinsically disordered proteins (IDPs) make up a large class of proteins that lack stable structures in solution, existing instead as dynamic conformational ensembles. To perform their biological functions, many IDPs bind to other proteins or nucleic acids. Although IDPs are unstructured in solution, when they interact with binding partners, they fold into defined three-dimensional structures via coupled binding-folding processes. Because they frequently underlie IDP function, the mechanisms of this coupled binding-folding process are of great interest. However, given the flexibility inherent to IDPs and the sparse populations of intermediate states, it is difficult to reveal binding-folding pathways at atomic resolution using experimental methods. Computer simulations are another tool for studying these pathways at high resolution. Accordingly, we have applied 40 μs of unbiased molecular dynamics simulations and Markov state modeling to map the complete binding-folding pathway of a model IDP, the 59-residue C-terminal portion of the DNA binding domain of Drosophila melanogaster transcription factor Brinker (BrkDBD). Our modeling indicates that BrkDBD binds to its cognate DNA and folds in ∼50 μs by an induced fit mechanism, acquiring most of its stable secondary and tertiary structure only after it reaches the final binding site on the DNA. The protein follows numerous pathways en route to its bound and folded conformation, occasionally becoming stuck in kinetic traps. Each binding-folding pathway involves weakly bound, increasingly folded intermediate states located at different sites on the DNA surface. These findings agree with experimental data and provide additional insight into the BrkDBD folding mechanism and kinetics.