Isotopologues are valuable vibrational probes that shift features in a vibrational spectrum while preserving the electronic structure of the molecule. We report the vibrational and electronic spectra of perdeuterated tryptophan in solution (L-Trp-d(5)), as Trp48-d(5) in azurin, and as the photogenerated neutral tryptophan radical, Trp48-d(5)(center dot), in azurin. The UV resonance Raman bands of the perdeuterated closed-shell tryptophan in solution and in azurin are lower in frequency relative to the protiated counterpart. The observed decrease in frequencies of L-Trp-d(5) bands relative to L-Trp-h(5) enables the analysis of vibrational markers of other amino acids, e.g., phenylalanine, that overlap with some modes of L-Trp-h(5). The Raman intensities vary between L-Trp-d(5) and L-Trp-h(5); these differences likely reflect modifications in normal mode composition upon perdeuteration. Analysis of the W3, W6, and W17 modes suggests that the W3 mode retains its utility as a conformational marker; however, the H-bond markers W6 and W17 appear to be less sensitive upon perdeuteration. The neutral tryptophan radical, Trp48-d(5)(center dot), was generated in azurin with a slightly lower radical quantum yield than for Trp48-h(5)(center dot). The visible resonance Raman spectrum of Trp48-d(5)(center dot) is different from that of Trp48-h(5)(center dot), especially in terms of relative intensities, and all assignable peaks decreased in frequency upon perdeuteration. The absorption and emission spectra of the perdeuterated closed-shell and radical species exhibited hypsochromic shifts of less than 1 nm relative to the protiated species. The data presented here indicate that L-Trp-d(5) is a valuable probe of vibrational structure, with minimal modification of photoreactivity and photophysics compared to L-Trp-h(5).