UV Resonance Raman Investigation of the Aqueous Solvation Dependence of Primary Amide Vibrations

Abstract

We investigated the normal mode composition and the aqueous solvation dependence of the primary amide vibrations of propanamide. Infrared, normal Raman, and UV resonance Raman (UVRR) spectroscopy were applied in conjunction with density functional theory (DFT) to assign the vibrations of crystalline propanamide. We examined the aqueous solvation dependence of the primary amide UVRR bands by measuring spectra in different acetonitrile/water mixtures. As previously observed in the UVRR spectra of N-methylacetamide, all of the resonance enhanced primary amide bands, except for the Amide I (AmI), show increased UVRR cross sections as the solvent becomes water-rich. These spectral trends are rationalized by a model wherein the hydrogen bonding and the high dielectric constant of water stabilizes the ground state dipolar − O -C=NH 2 + resonance structure over the neutral O=C-NH 2 resonance structure. Thus, vibrations with large C-N stretching show increased UVRR cross sections because the C-N displacement between the electronic ground and excited state increases along the C-N bond. In contrast, vibrations dominated by C=O stretching, such as the AmI, show a decreased displacement between the electronic ground and excited state, which result in a decreased UVRR cross section upon aqueous solvation. The UVRR primary amide vibrations can be used as sensitive spectroscopic markers to study the local dielectric constant and hydrogen bonding environments of the primary amide side chains of glutamine (Gln) and asparagine (Asn). Graphical Abstract *Corresponding Author: asher@pitt.edu. NotesThe authors declare no competing financial interest. Tables S1-S6 and Figures S1-S3. Description of X-ray crystallographic methods to determine structure of propanamide and UVRR spectral deconvolution methods. This material is available free of charge via the Internet at http://pubs.acs.org/. Supporting Information

Glutamine and Asparagine Side Chain Hyperconjugation-Induced Structurally Sensitive Vibrations

“…On the basis of our normal mode calculations of Gln, butyramide (Supporting Information Table S14), and propanamide, 28 we conclude that N ε H 2 rocking, C δ –N ε stretching, and C β –C γ stretching define the AmIII P vibration. However, depending on the OCCC dihedral angle, other motions such as C β H 2 twisting and C α –C β stretching can contribute to this vibration.…”

“…We employ DFT calculations that use a more modern functional (M06-2X) than that of Ramirez and co-workers. These assignments build off of our previous, detailed assignment of propanamide, 28 a model for the side chains of Gln and Asn. Our assignments of l -Gln in H 2 O and D 2 O are shown in Tables 1 and 2, respectively.…”

“…28 In water, the AmIII P band of l -Gln is located at ∼1110 cm −1 , as compared with ∼1097 cm −1 in the solid-state. On the basis of Rhys et al's neutron diffraction study, 58 the solution-state equilibrium structure of l -Gln in water does not appear to differ significantly from the single known l -Gln crystal structure.…”

“…17–28 Deep UV excitation (∼200 nm) selectively resonance enhances secondary and primary amide vibrations. 29,30 Previous investigations of secondary amide vibrations have developed a detailed understanding of the UVRR spectral dependence on the peptide bond structure and its hydrogen bonding.…”

Ultraviolet resonance Raman spectroscopic markers for protein structure and dynamics

“…Using UVRR, Punihaole et al investigated the aqueous solvation dependence of the primary amide vibrations of propanamide, a small primary amide compound model for Gln and Asn side chains [33]. They discovered that almost all the bands in the UVRR spectrum increase in both intensity and frequency with increasing water solvation.…”