Application of Density Functional Theory Calculations to Favoenzymes

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Density functional theory was applied to acyl-CoA dehydrogenase(ACD) and the calculated results were discussed with reference to the experimental ones. Especially, we focused on the CT interaction observed in the complex between ACD and substrate analog. 3-Thiaacyl-CoA is known to be a transition-state analog, which is transferred to an anionic form through an α-proton abstraction process by a catalytic base glutamate and interacts with oxidized flavin in the active site. The common α-proton abstraction process is caused in the dehydrogenation of substrate, thus the CT interaction with 3-thiaacyl-CoA is expected to reflect the initial stage of dehydrogenation reaction.The calculation model was simplified as the complex of lumiflavin and ethylthioester of 3-thiabutanoic acid, and the model was energetically optimized using the density functional of B3LYP/6-311G(d). The geometry and orientation of the two molecules was optimized to reach a structure that was quite similar to the X-ray structure reported. This represents that the spatial arrangement of the CT complex is determinant due to electrostatic interaction between an oxidized flavin and an anionic 3-thiaacyl chain. Electronic absorption spectrum was also simulated in this molecular geometry on the same basis set. The calculations reproduced a CT band at long-wavelength region.In this project, we also have developed the methodology on difference infrared absorption(IR) spectra to selectively observe a vibrational mode in the enzyme complex. This method utilizes the synthetic compounds(flavins or ligands) labeled with stable isotopes, e.g.13C,15N or 18O. Calculated vibrational frequencies and infrared intensities showed to reproduce the experimental ones. This method is highly expected to be applicable to the other enzyme systems to investigate the molecular interactions in the active sites.