DNA binding redistributes activation domain ensemble and accessibility in pioneer factor Sox2.

More than 1600 human transcription factors orchestrate the transcriptional machinery to control gene expression and cell fate. Their function is conveyed through intrinsically disordered regions (IDRs) containing activation or repression domains but lacking quantitative structural ensemble models prevents their mechanistic decoding. Here we integrate single-molecule FRET and NMR spectroscopy with molecular simulations showing that DNA binding can lead to complex changes in the IDR ensemble and accessibility. The C-terminal IDR of pioneer factor Sox2 is highly disordered but its conformational dynamics are guided by weak and dynamic charge interactions with the folded DNA binding domain. Both DNA and nucleosome binding induce major rearrangements in the IDR ensemble without affecting DNA binding affinity. Remarkably, interdomain interactions are redistributed in complex with DNA leading to variable exposure of two activation domains critical for transcription. Charged intramolecular interactions allowing for dynamic redistributions may be common in transcription factors and necessary for sensitive tuning of structural ensembles.

Multilayered allosteric modulation of coupled folding and binding by phosphorylation, peptidyl-prolyl cis/trans isomerization, and diversity of interaction partners.

Intrinsically disordered proteins (IDPs) play key roles in cellular regulation, including signal transduction, transcription, and cell-cycle control. Accordingly, IDPs can commonly interact with numerous different target proteins, and their interaction networks are expected to be highly regulated. However, many of the underlying regulatory mechanisms have remained unclear. Here, we examine the representative case of the nuclear coactivator binding domain (NCBD) of the large multidomain protein CBP, a hub in transcriptional regulation, and the interaction with several of its binding partners. Single-molecule Förster resonance energy transfer measurements show that phosphorylation of NCBD reduces its binding affinity, with effects that vary depending on the binding partner and the site and number of modifications. The complexity of the interaction is further increased by the dependence of the affinities on peptidyl-prolyl cis/trans isomerization in NCBD. Overall, our results reveal the potential for allosteric regulation on at least three levels: the different affinities of NCBD for its different binding partners, the differential modulation of these affinities by phosphorylation, and the effect of peptidyl-prolyl cis/trans isomerization on binding.

Simulating Polyproline II-Helix-Rich Peptides with the Latest Kirkwood-Buff Force Field: A Direct Comparison with AMBER and CHARMM.

We simulated the dynamics of a set of peptides characterized by ensembles rich in PPII-helical content, to assess the ability of the most recent Kirkwood-Buff force field (KBFF20) to sample this conformational peculiarity. KBFF has been previously shown to capably reproduce experimental dimensions of disordered proteins, while being limited in confidently sampling structured proteins. Further development of the force field bridged this gap. It is however still unknown what are the main differences between KBFF and AMBER/CHARMM force fields. A direct comparison is now possible as both AMBER/CHARMM force fields have been used to sample peptides rich in PPII-helical content. We found that KBFF20 samples' PPII-helical content qualitatively matches both AMBER and CHARMM force fields, with the main difference being the KBFF ability to populate the αR region of the Ramachandran plot in the set of simulated peptides. Overall, KBFF20 is a well-balanced force field, able to sample the dynamics of both structured and unstructured proteins.

Protein intrinsic disorder on a dynamic nucleosomal landscape.

The complex nucleoprotein landscape of the eukaryotic cell nucleus is rich in dynamic proteins that lack a stable three-dimensional structure. Many of these intrinsically disordered proteins operate directly on the first fundamental level of genome compaction: the nucleosome. Here we give an overview of how disordered interactions with and within nucleosomes shape the dynamics, architecture, and epigenetic regulation of the genetic material, controlling cellular transcription patterns. We highlight experimental and computational challenges in the study of protein disorder and illustrate how integrative approaches are increasingly unveiling the fine details of nuclear interaction networks. We finally dissect sequence properties encoded in disordered regions and assess common features of disordered nucleosome-binding proteins. As drivers of many critical biological processes, disordered proteins are integral to a comprehensive molecular view of the dynamic nuclear milieu.