Deciphering Infrared Difference Spectra of Bovine Rhodopsin in the Amide I Region by Localized Anharmonic Vibrational Calculations.

Vibrational quasi-degenerate perturbation theory (VQDPT2) based on localized vibrational modes is combined with quantum mechanics/molecular mechanics (QM/MM) calculations to investigate the amide bands of bovine rhodopsin in the dark (inactive) and Meta II (active) states. A novel localization scheme has been developed for this purpose: (1) the protein is partitioned into individual amide-group subsystems, (2) the vibrational spectrum of each amide group is calculated by QM/MM and VQDPT2, and (3) the total spectrum is constructed by summing the individual local amide spectra. Our approach reproduces the experimental IR difference spectrum upon photoactivation across the amide I, II, and III regions, demonstrating the importance of accounting for anharmonicity. Detailed analysis has revealed that the amide I band signals are not localized to a single domain; rather, the higher-wavenumber shifts are primarily driven by helical regions, while lower-wavenumber shifts stem from loop regions. We identified specific residues, such as Thr118 and Trp265, whose significant frequency shifts are coupled to local structural rearrangements. This subsystem-based anharmonic vibrational approach provides a powerful tool for the microscopic interpretation of vibrational spectra in large biomolecular systems.