Hidden electronic excited state of enhanced green fluorescent protein (eGFP) by new multiplex two-photon absorption spectroscopy

PHYS 27

Haruko Hosoi, haru@riken.jp1, Shoichi Yamaguchi1, Hideaki Mizuno2, Atsushi Miyawaki2, and Tahei Tahara1. (1) Molecular Spectroscopy Laboratory, RIKEN (The Institute of Physical and Chemical Research), 2-1 Hirosawa, Wako, 351-0198, Japan, (2) Brain Science Institute, RIKEN (The Institute of Physical and Chemical Research), 2-1 Hirosawa, Wako, 351-0198
We have newly developed non-degenerate two-photon absorption (TPA) spectroscopy, which enables us to measure precise TPA spectra for a broad wavelength range, utilizing narrow-band and broad-band femtosecond pulses with multichannel detection. We applied this technique to study the electronic structure of enhanced green fluorescent protein (eGFP), which is the most widely used GFP as a fluorescent probe for bioimaging. The observed TPA spectrum is significantly blue-shifted compared with its one-photon absorption spectrum although the chromophore of eGFP has no inversion symmetry. The result indicates the existence of a “hidden” excited state in the vicinity of the lowest excited singlet state. We conclude that this is the origin of the discrepancy between the one-photon and two-photon excitation spectra of eGFP, which is well known in the field of biology.
 

Optical Probes of Dynamics in Complex Environments
8:00 AM-12:00 PM, Sunday, April 6, 2008 Morial Convention Center -- Rm. 344, Oral

Division of Physical Chemistry

The 235th ACS National Meeting, New Orleans, LA, April 6-10, 2008