Glutamine and Asparagine Side Chain Hyperconjugation-Induced Structurally Sensitive Vibrations

We identified vibrational spectral marker bands that sensitively report on the side chain structures of glutamine (Gln) and asparagine (Asn). Density functional theory (DFT) calculations indicate that the Amide III^P (AmIII^P) vibrations of Gln and Asn depend cosinusoidally on their side chain OCCC dihedral angles (the x₃ and x₂ angles of Gln and Asn, respectively). We use UV resonance Raman (UVRR) and visible Raman spectroscopy to experimentally correlate the AmIII^P Raman band frequency to the primary amide OCCC dihedral angle. The AmIII^P structural sensitivity derives from the Gln (Asn) C_β-C (C_α-C_β) stretching component of the vibration. The C_β-C (C_α-C_β) bond length inversely correlates with the AmIII^P band frequency. As the C_β-C (C_α-C_β) bond length decreases, its stretching force constant increases, which results in an upshift in the AmIII^P frequency. The C_β-C (C_α-C_β) bond length dependence on the x₃ (x₂) dihedral angle results from hyperconjugation between the CO (CO) ∗ and C_β-C (C_α-C_β) orbitals. Using a Protein Data Bank library, we show that the x₃ and x₂ dihedral angles of Gln and Asn depend on the peptide backbone Ramachandran angles. We demonstrate that the inhomogeneously broadened AmIII^P band line shapes can be used to calculate the x₃ and x₂ angle distributions of peptides. The spectral correlations determined in this study enable important new insights into protein structure in solution, and in Gln- and Asn-rich amyloid-like fibrils and prions.