Vibrationally Resolved Photoabsorption Spectroscopy of Red Fluorescent Protein Chromophore Anions

Photoabsorption studies of red fluorescent protein chromophore anions have been performed at the ELISA electrostatic heavy-ion storage ring. The broad absorption band due to electronic excitation of the chromophores is tuned to a longer wavelength (redshifted) by extending the electronic conjugation of the molecule. A clear vibrational progression is resolved with $E_{\mathrm{v}\mathrm{i}\mathrm{b}}\sim 380$ and $520{\mathrm{c}\mathrm{m}}^{-1}$ for two different forms of the chromophore. The vibrational modes correspond to collective motions of the entire molecular structure. It is argued that the excited electronic state has an equilibrium configuration far from that of the electronic ground state, i.e., poor Franck Condon overlap.

Figure 1

Structure of model DsRed chromophores in which the GFP chromophore core has been extended by one [DsRed(1)] and two [DsRed(2)] ethylenic bonds.

Figure 2

The electrostatic ion storage ring ELISA equipped with an electrospray ion source, a pulsed laser, and a detector for neutral products.

Figure 3

Yield of neutrals detected as a function of time and laser wavelength for DsRed(2) anions.

Figure 4

The time-integrated absorption spectra of DsRed(1) and DsRed(2) anions. Shown by bars are vibrational structures, the energies of which are shown to the right. The vibrational spacing is $382{\mathrm{c}\mathrm{m}}^{-1}$ for DsRed(1) and $518{\mathrm{c}\mathrm{m}}^{-1}$ for DsRed(2) with an estimated uncertainty of $10{\mathrm{c}\mathrm{m}}^{-1}$.