Optical microresonator modifies the efficiency of the fluorescence resonance energy transfer in the autofluorescent protein DsRed

Abstract

We investigate experimentally the modifications of the fluorescence properties of the bichromophoric fluorescent resonance energy transfer (FRET) system DsRed imposed by optical confinement. The confinement-condition is realized by a novel Λ/2-microresonator that modifies the local photonic mode density in the vicinity of the proteins while maintaining a physiological environment for the embedded biological molecules. The experimental ratio of the fluorescence intensities and lifetimes, respectively, of donor and acceptor chromophores varies by up to a one order of magnitude as we vary the mirror spacing of the microresonator with nanometer-precision. Since these ratios determine the FRET efficiency, we modify the yield of the excited state energy transfer in rigidly coupled FRET pairs without chemically or physically perturbating the chromophoric subunits. We show that the microresonator- controlled inhibition of the acceptor fluorescence results in a loss of transfer efficiency of excited state energy from donor to acceptor, an effect that enables the spectral isolation and efficient observation of donor chromophores both in DsRed ensembles and on the single protein level. This constitutes an important application of microcavity-enhanced single molecule spectroscopy of biological systems and shows the potential of optical confinement for applications in nano-biophotonics. ©2009 SPIE.