The conformational ensemble of OPN thus contains both, cooperatively folded and unfolded, extended conformations. Additionally, EPR and NMR (PRE) experiments under high NaCl concentrations showed that not only hydrophobic interactions contribute to the OPN’s structural stability, but also electrostatics play a crucial role in the stabilization of compact structures of OPN in solution [46]. The surprisingly Epacadostat nmr detailed picture of the conformational ensemble of OPN obtained by this novel approach indicates valuable applications

to studies of structural dynamics of IDPs. IDPs are characterized by rugged energy landscapes devoid of distinct energy barriers and therefore display significant structural plasticity and undergo large structural rearrangements. A comprehensive characterization of the solution structures of IDPs thus requires studies of conformational dynamics. NMR spectroscopy is destined for these studies and a

plethora of different experiments are available providing detailed information about motional dynamics on different time scales. Fast (ps–ns) time scale motions are probed by 15N spin relaxation experiments (15N-T1,T2 and 15N–1HN NOEs) and analyzed using well-established theoretical frameworks (e.g. model-free formalism [47]). Slower motions occurring on μs–ms time scales are investigated by CPMG-type schemes introduced decades ago and turned into a powerful experimental methodology applicable even to very selleck chemical large molecular weight systems by Kay and co-workers [48]. The particular uniqueness of NMR spin relaxation measurements is the fact that detailed information about internal motions can be discerned. In case of globular, stably folded proteins the analysis relies on distinctly nearly different correlation times describing overall tumbling and internal motions. In case of IDPs this clear-cut separation is no longer valid and thus hampers the application of this approach.

A similar situation was encountered in RNA NMR studies. In order to overcome this limitation the group of Al-Hashimi developed an elegant domain elongation strategy to effectively decouple internal motions from overall tumbling [49]. In a similar way we adapted this strategy to relaxation studies of IDPs. As a first example we studied the internal dynamics of OPN using dimeric Myc/Max protein complex for domain elongation. The crystal structure of Myc/Max revealed a four helical bundle structure with significant overall anisotropy. OPN was covalently attached to Myc via a Bismaleimide linker (C54@OPN–C34@Myc). 15N NMR relaxation data obtained for the OPN-Myc/Max complex were compared with data obtained from isolated OPN (Fig. 10).