Predicting the optical properties of fluorescence probes in a protein matrix: a 2-rhodamines-labeled biosensor for inorganic phosphate

Abstract

Fluorescence probes are routinely used to investigate the structural basis of biochemical processes in vitro and in vivo, yet they are poorly understood. Here we will calculate the optical properties of rhodamine-based probes attached to the phosphate binding protein, which are used as inorganic phosphate sensors. Binding of P_i to the protein provokes structural rearrangements of the protein. This is associated with a drastic change in the absorption and emission spectra of the rhodamines, along with an enhancement in the emission intensity of about 18 folds. The structural facets on which this process is based are not clear. Here we will use a variety of computational approaches, including force field based metadynamics as well as quantum mechanical techniques such as TDDFT/MM and CASPT2. Our calculations will be validated against experiments performed by the experimental group of the team. This work will establish the molecular basis of fluorescence and absorption spectral shifts upon ligand-used conformational changes and may be used as a general protocol in a variety of biophysical applications.