with poly-alanine reference structures, nor in the weighted average of the 20 cluster center structures (Fig. ). Finally, we extended this side chain rotamer approach to the first four cluster center structures to verify how one specific side chain configuration of the peptide affects the simulated spectra (ESI Fig. ). While for IR each side chain configuration yields a specific spectrum, but with the same overall characteristic of the averaged one (ESI Fig. ), Raman is found more sensitive to the signals arising from the side chains and their orientation (ESI Fig. ). The ROA spectra are almost entirely determined by a specific side chain configuration (all χ-angles of all residues, ESI Fig. ). Only the positive amide I signal is conserved in all calculations irrespective of the side chain conformation, and is thus a robust marker of the backbone conformation. In conclusion, this clearly demonstrates the need for thorough averaging over many conformations of both the backbone and side chain groups, as experimental ROA spectra show distinct patterns depending on the backbone conformation.