Resolving Electronic Transitions in Synthetic Fluorescent Protein Chromophores by Magnetic Circular Dichroism

Abstract

The detailed electronic structures of fluorescent chromophores are important for their use in imaging of living cells. A series of green fluorescent protein chromophore derivatives is examined by magnetic circular dichroism (MCD) spectroscopy, which allows the resolution of more bands than plain absorption and fluorescence. Observed spectral patterns are rationalized with the aid of time-dependent density functional theory (TDDFT) computations and the sum-over-state (SOS) formalism, which also reveals a significant dependence of MCD intensities on chromophore conformation. The combination of organic and theoretical chemistry with spectroscopic techniques also appears useful in the rational design of fluorescence labels and understanding of the chromophore's properties. For example, the absorption threshold can be heavily affected by substitution on the phenyl ring but not much on the five-member ring, and methoxy groups can be used to further tune the electronic levels.

On the magnetic circular dichroism of benzene. A density-functional study

“…Degenerate electronic levels (Faraday A-terms) were also included, as a limit of close-lying states. 53 The Kaufmann's basis set optimized to represent Rydberg states 54 was placed at the center of benzene molecule; by default, orbitals up to the main quantum number (n) of 7 and the orbital quantum number (l) of 1 were taken. We indicate this by the letter "R," e.g., B3LYP/6-311++G**/R (or just B3LYP/R) calculation was performed with the standard 6-311++G** basis set augmented by the diffuse functions.…”

Vibrational Structure in Magnetic Circular Dichroism Spectra of Polycyclic Aromatic Hydrocarbons

“…Note that enantiomers provide the same MCD: the coordinate inversion changes only signs of the electric dipole moments or gradients, which always multiply to unity in the expression for B. Degenerate electronic levels, conventionally giving rise to Faraday A-terms, were included as a limit of close-lying states, 47 i.e., by replacing expressions Alternatively to TDDFT, single-and multi-state complete active space second-order perturbation theories (SS-CASPT2 27 and MS-CASPT2, 48 respectively) were used to obtain the ground and excited electronic states. MCD intensities were calculated using the method of Ganyushin and Neese.…”