Reaction-Based Genetically Encoded Fluorescent Hydrogen Sulfide Sensors

Abstract

The detection of hydrogen sulfide (H(2)S), a toxic gas and an important biological signaling molecule, has been a long-time challenge. Here we report genetically encoded fluorescent protein (FP)-based probes that can selectively detect H(2)S. By expanding the genetic codes of E. coli and mammalian cells, FP chromophores were modified with the sulfide-reactive azide functional group. These structurally modified chromophores were selectively reduced by H(2)S, resulting in sensitive fluorescence enhancement detectable by spectroscopic and microscopic techniques. Exploration of a circularly permuted FP led to an improved sensor with faster responses, and the feasibility of using such a genetically encoded probe to monitor H(2)S in living mammalian cells has also been demonstrated.

Highly sensitive sulphide mapping in live cells by kinetic spectral analysis of single Au-Ag core-shell nanoparticles

“…During the past several years, several research groups have developed sulphide-responsive fluorescent organic molecules to detect H 2 S in live cells and have achieved selective intracellular sulphide imaging with limits of detection of 1-10 mM (ref. [23][24][25][26][27][28]. Although most publications suggest that the average endogenous H 2 S level is in the mM range, much lower sulphide concentrations have been reported 29 .…”

Expanding the chemistry of fluorescent protein biosensors through genetic incorporation of unnatural amino acids

“…14 UAA*s ( Fig. 2) that are successfully applied to the biosensor design include 3,4-dihydroxy-L-phenylalanine (L-DOPA; metal ion sensor), 10,45 2-amino-3-(8-hydroxyquinolin-5-yl)propanoic acid (HqAla; metal ion sensor), 13 para-borono-L-phenylalanine (pBoF; H 2 O 2 sensor), 11,44 para-azido-L-phenylalanine (pAzF; H 2 S sensor), 12,45 and o-nitrobenzyl-O-tyrosine (ONBY, light-responsive FP). 14,46 49 HqAla, 13 and bipyridyl-L-alanine (BpA), 50 into fluorescent proteins.…”

“…A reaction-based FP sensor for H 2 S was also constructed by Chen and co-workers. 12 As shown in Fig. 5, the chromophore-forming Tyr67 residue of teal fluorescent protein (mTFP1) was replaced with pAzF.…”

“…An intriguing question is whether the introduction of non-canonical functional groups with novel physical, chemical, and biological properties through genetic incorporation of unnatural amino acids (UAA*s) into FPs could further diversify the proteins' structure and function and lead to novel designs of FP biosensors. The combination of FP engineering and UAA* mutagenesis methods will provide a new frontier for the design and construction of FP biosensors, [10][11][12][13][14] which is the main focus of this review. The intention of this review is to attract more attention towards this emerging research field and to provoke creative designs and innovative utilization of FP biosensors containing unnatural elements for biological studies in living systems.…”

Simultaneous Imaging of Ribonucleic Acid and Hydrogen Sulfide in Living Systems with Distinct Fluorescence Signals Using a Single Fluorescent Probe

“…More recently, the well‐designed synthetic fluorescent nucleic acids dyes, including commercially available nucleic acids dyes, have been employed in bioimaging fields 10. Over the past few years, a wide variety of fluorescent H 2 S probes have been constructed 11. However, there have been no reports on probes for simultaneous imaging RNA and H 2 S in living systems.…”