Turning on and off photoinduced electron transfer in fluorescent proteins by pi-stacking, halide binding, and Tyr145 mutations
Photoinduced electron transfer in fluorescent proteins (FP) from the green fluorescent protein (GFP) family is a novel phenomenon [Bogdanov et al, 2009]. Depending on the task at hand, it is regarded either as an asset facilitating new applications or as a nuisance leading to the loss of optical output. In many FPs photooxidation of the anionic chromophore results in a green-to-red photoconversion called oxidative redding. We discovered that yellow FPs (YFPs) that are closely related to GFP and share the same anionic chromophore do not undergo redding. The redding is restored in some YFPs upon halide binding. Detailed calculations of the energetics of one-electron oxidation process and possible ET pathways suggested that excited-state ET proceeds predominantly through a hopping mechanism via the Tyr145 residue. In YFPs, the pi-stacking of the chromophore with Tyr203 reduces the electron donating ability of the chromophore and suppresses the main ET channel. The halide binding restores favorable energetics by upsetting the pi-stacking and by modifying local electrostatic field. The theoretical predictions were validated by point mutations that confirmed that Tyr145 is a key residue controlling ET. Substitution of Tyr145 by less efficient electron acceptors resulted in mutants with extremely high photostabilities. We believe that this strategy - calculation and further disruption of ET pathways - may represent a new widely applicable approach for enhancing photostability of FPs.