*4.3. Restrained Modeling*

The set of the experimentally determined backbone and Cβ chemical shifts were input to run restrained MD modeling by means of the CS-ROSETTA server [19]. The chemical shifts represent experimental information that is employed to restrain the backbone dihedral angles *ϕ* and *ψ* by means of a pseudopotential term that introduces an energy penalty upon violation [19]. The same energy-penalty-driven approach was employed to calculate the structure based on the inter-proton distances obtained from the 2D and 3D NOESY spectra. The NOE-restrained structure determination was handled by means of the

software suite PONDEROSA-C/S, using PONDEROSA-X refinement by which automated database-assisted NOE assignment is done (AUDANA algorithm) [50]. Experimentally determined chemical shift assignments for backbone, sidechain, and aromatic residues were input to automatically assign the 3D <sup>15</sup>N-1H NOESY HSQC spectrum, and aliphatic and aromatic 3D <sup>13</sup>C-1H NOESY HSQC spectra and calculate the structure as per the above procedure. Automated NOE-assignments were manually checked to remove ambiguous assignments and to add additional constraints. NOE intensities were considered only qualitatively as strong, medium and weak, corresponding to upper limit distances of 0.25, 0.35 and 0.5 nm, respectively.
