Vibrational Raman optical activity (ROA) spectroscopy [1] measures the difference in Raman scattering intensity of chiral molecules in right-and left-polarized incident light, and allows one to determine the structure and absolute configuration of biomolecules in aqueous solution.[2] While most bands in the ROA spectra of proteins are assigned to the peptide backbone and can provide information on secondary structure elements (see, eg, ref.[3] for experimental work and refs.[4, 5] for theoretical studies), some bands are related to the conformation of specific side chains. By comparing the ROA backscattering spectra of different viral coat proteins, Blanch et al. suggested [6] that a ROA band at% 1550 cmÀ1, which is assigned to the W3 vibration of the indole ring in tryptophan, can be used to determine the absolute stereochemistry of the tryptophan side chain. While it had been shown earlier in an analysis of the Raman spectra of different crystalline tryptophan derivatives [7] that the wavenumber of this W3 vibration correlates with the magnitude of the c2, 1 torsion angle (see Figure 1 for the definition of this angle), they inferred that its sign can be deduced from the sign of the corresponding ROA band. From the positive W3 ROA signal of hen lysozyme in solution and the observation that its crystal structure shows a positive sign of c2, 1 for four of the six tryptophan residues, they concluded that a positive W3 ROA signal corresponds to a positive sign of c2, 1. Additional support for this assignment is given by an observed increase of the magnitude of the W3 ROA signal for a complex of hen lysozyme with the trimer of N-acetylglucosamide in which, according to its crystal structure, five out of six tryptophans show a positive sign of c2, 1.

However, there are a number of uncertainties in this assignment which could not be resolved by experiment. The sign and the magnitude of the W3 ROA signal might depend not only on c2, 1 (Blanch et al. assumed a sinc2, 1 dependence of the magnitude of the ROA intensity), but could also be influenced by other structural parameters, such as the χ1 torsion angle. This would complicate the comparison of the W3 ROA signal observed in hen lysozyme to the crystal structure, which contains six tryptophan residues in different conformations. Furthermore, it is not certain that the conformation of the tryptophan side chains in solution is the same as in the crystal. To address these issues, we have investigated the influence of the torsion angles c2, 1 and c1 on the intensity of the W3 ROA signal in tryptophan by performing density functional theory (DFT) calculations. These calculations allow us to study the effect of the different structural changes on the ROA spectrum in a controlled manner and further make it possible to analyze the origin of the ROA signals of interest. As model system, we chose N-acetyl-(S)-tryptophan-Nо-methylamide (see Figure 1), which is the simplest model that, in addition to the tryptophan side chain, also contains the main structural features of the peptide backbone. This structural model allows for an extrapolation of the results to extended peptide chains and proteins because it is reasonable to assume that the normal mode responsible for the W3 signal is localized on the tryptophan side chain and does not couple with normal modes on other parts of the protein. We further expect that the ROA intensity is not significantly influenced by contributions of other chiral residues that are not included in the model to the total wavefunction.