UV resonance Raman finds peptide bond-Arg side chain electronic interactions

Abstract

We measured the UV resonance Raman excitation profiles and Raman depolarization ratios of the arginine (Arg) vibrations of the amino acid monomer as well as Arg in the 21-residue predominantly alanine peptide AAAAA(AAARA)(3)A (AP) between 194 and 218 nm. Excitation within the π → π* peptide bond electronic transitions result in UVRR spectra dominated by amide peptide bond vibrations. The Raman cross sections and excitation profiles indicate that the Arg side chain electronic transitions mix with the AP peptide bond electronic transitions. The Arg Raman bands in AP exhibit Raman excitation profiles similar to those of the amide bands in AP which are conformation specific. These Arg excitation profiles distinctly differ from the Arg monomer. The Raman depolarization ratios of Arg in monomeric solution are quite simple with ρ = 0.33 indicating enhancement by a single electronic transition. In contrast, we see very complex depolarization ratios of Arg in AP that indicate that the Arg residues are resonance enhanced by multiple electronic transitions.

Figure 1

AP conformations (upper panel) and structure (lower panel). AP is predominantly α-helical at low temperatures, and PPII-like at high temperatures.

Figure 2

UV absorption spectrum of arg at pH 7 and 25 °C; UV absorption spectra of AP at 5 °C, where it is predominantly α-helical, and at 60 °C, where it is predominantly in a PPII-like conformation.

Figure 3

UVRR spectra of Arg at 25° C between 218 and 194 nm. The spectra were collected for 15 min at each excitation wavelength. The spectral resolution varies between 5.9 cm⁻¹ at 218 nm and 7 cm⁻¹ at 194 nm. All spectra were normalized to the ClO₄⁻ internal standard (not shown). Inset: Arg structure.

Figure 4

UVRR spectra of AP (pH 7) at 60 °C and 5 °C excited at 204 nm. The spectra were collected for 15 min at each excitation wavelength. The Arg bands in the AP

Figure 5

a) Absolute Raman cross section (mbarn·mol⁻¹·sr⁻¹) excitation profiles between 194 to 218 nm of Arg in solution at 25 °C. Also shown is the Arg absorption spectrum. b) Absolute Raman cross section excitation profiles (mbarn·Arg residue⁻¹·sr⁻¹) of Arg vibrations in the PPII-like conformation of AP, and the AP AmII Raman cross section excitation profile (mbarn·peptide bond⁻¹·sr⁻¹) of the PPII-like conformation. c) Absolute Raman cross section excitation profiles (mbarn·Arg residue⁻¹·sr⁻¹) of the Arg vibrations of the α-helix conformation of AP, and the α-helix conformation AP peptide bond AmII Raman cross section excitation profile (mbarn·peptide bond⁻¹·sr⁻¹).

Figure 6

Overlapped (mbarn · peptide bond⁻¹ · sr⁻¹) cross sections of (a) the AP AmII vibration and Arg 1633 cm⁻¹ band in AP PPII-like conformation; (b) AP AmII vibration and Arg 1633 cm⁻¹ band in AP α-helix conformation.

Figure 7

Depolarization ratios of Arg in water. These ρ = 0.33 values indicate the occurrence of a single electronic transition

Figure 8

Depolarization ratios of the Arg vibrations and of two AP amide vibrations in the PPII-like conformation of AP. Single electronic transitions ρ = 0.33 (aqua), totally non-symmetric transitions ρ = 0.75 (purple) and totally symmetric transitions, ρ = 0.125 (pink) ρ values are indicated by solid lines.

Figure 9

Depolarization ratios for the Arg vibrations and for two AP amide vibrations in the α-helix conformation of AP. Single electronic transitions ρ = 0.33 (aqua), totally non-symmetric transitions ρ = 0.75 (purple) and totally symmetric transitions, ρ = 0.125 (pink) are indicated by solid lines.