Decoding Order and Disorder in Proteins by NMR Spectroscopy.

Proteins often have a complex architecture, consisting of both globular ordered domains and intrinsically disordered regions (IDRs). These multidomain proteins pose challenges for traditional structural biology techniques. One major difficulty arises from the dynamic and flexible nature of IDRs, which lack a stable three-dimensional structure. Indeed, this feature further complicates the application of traditional structural biology techniques. Characterizing these systems is typically simplified by isolating individual domains, which can provide valuable insights into the structure and function of specific regions. However, this approach overlooks the interactions and regulatory mechanisms that occur between domains. To capture the full functional and structural complexity of multidomain proteins, it is crucial to study larger constructs. In this study, we focused on the CREB binding protein (CBP), a pivotal protein involved in numerous cellular processes. CBP is characterized by its modular structure, featuring alternating globular domains and IDRs. We specifically examined the TAZ4 construct, encompassing the TAZ2 globular domain and the ID4 flexible linker region. To characterize this multidomain system, we designed NMR experiments that take advantage of the dynamic differences between the two domains to obtain 2D and 3D spectra enabling the selection of the signals based on their nuclear relaxation properties. These experiments allowed the sequence-specific assignment of the TAZ4 construct to be extended revealing a crosstalk between the disordered region and the globular domain.