Near-UV Circular Dichroism and UV Resonance Raman Spectra of Tryptophan Residues as a Structural Marker of Proteins

Abstract

Near-UV circular dichroism (CD) and UV resonance Raman (UVRR) spectra of L-tryptophan (Trp), its derivatives, and indole-C3 derivatives were investigated to utilize Trp signals of proteins as a structural marker. CD spectra of Trp are classified into four types: Free L-Trp gives type II (around 270 nm, L(a) transition), while L-Trp in proteins generally yields type I (around 280-290 nm, L(b) transition) often with vibronic structures. All the indole-C3 derivatives except for L-Trp gave no CD bands for L(a) and L(b) transitions, indicating that the asymmetric carbon (Cα) connected through C3-Cβ is essential to appearance of CD. We demonstrate here that the type of CD spectra is determined by a condition of the amino group of Trp; it was changed from type II to type I by the modification of the amino group. In contrast, the modification of the carboxyl group of L-Trp had little effects on a CD spectrum. The 229 nm excited UVRR spectra were almost the same between L-Trp and indole-C3 derivatives. Comparison of CD and UVRR spectra of Trp residues in proteins suggested that mainly the W17 (possibly together with W16) mode contributes to the characteristic vibronic coupling of L(b) transition. Both UVRR and CD spectra of L-Trp were influenced by protonation of amino and/or carboxyl groups, but those changes were distinguished from hydrogen bonding effects at N1H of indole. It is likely that these protonations are communicated to indole through σ-bonds containing Cα and thus influence both chirality of L(a) and L(b) transitions and properties of the Bb excited state.