Discovery and Characterization of a Naturally Occurring, Turn-On Yellow Fluorescent Protein Sensor for Chloride.

Abstract

Fluorescent proteins have been extensively engineered and applied as optical indicators for chloride in a variety of biological contexts. Surprisingly, given the biodiversity of fluorescent proteins, a naturally occurring chloride sensor has not been reported to date. Here, we present the identification and spectroscopic characterization of the yellow fluorescent protein from the jellyfish Phialidium sp . (phiYFP), a rare example of a naturally occurring, excitation ratiometric, and turn-on fluorescent protein sensor for chloride. Our results show that chloride binding tunes the p Ka of the chromophore Y66 and shifts the equilibrium from the fluorescent phenolate form to the weakly fluorescent phenol form. The latter likely undergoes excited-state proton transfer to generate a turn-on fluorescence response that is pH-dependent. Moreover, anion selectivity and mutagenesis in the chloride binding pocket provide additional evidence for the proposed chloride sensing mechanism. Given these properties, we anticipate that phiYFP, with further engineering, could be a new tool for imaging cellular chloride dynamics.