A Diselenide Turn‐On Fluorescent Probe for the Detection of Thioredoxin Reductase

Abstract

We report the first diselenide‐based probe for the selective detection of thioredoxin reductase (TrxR), an enzyme commonly overexpressed in melanomas. The probe design involves conjugation of a seminaphthorhodafluor dye with a diselenide moiety. TrxR reduces the diselenide bond, triggering a fluorescence turn‐on response of the probe. Kinetic studies reveal favorable binding of the probe with TrxR with a Michaelis–Menten constant (Km) of 15.89 μm. Computational docking simulations predict a greater binding affinity to the TrxR active site in comparison to its disulfide analogue. In vitro imaging studies further confirmed the diselenide probe exhibited improved signaling of TrxR activity compared to the disulfide analogue.

Thioredoxin reductase-triggered fluorogenic donor of hydrogen sulfide: a model study with a symmetrical organopolysulfide probe with turn-on near-infrared fluorescent emission

“…Indeed, this is not surprising as (a) the structure of DCI-PS or DCI-DS highly resembles the structures of reported fluorogenic probes of TrxR having cyclic disulfide ( TRFS-green and FAST-TRFS ) or acyclic diselenide unit (probe 1a ) (Fig. 1b); 28–30 (b) the nucleophilicity of the selenol group of reduced TrxR must be much higher than that of the thiol group of GSH. However, unlike other related antioxidant proteins such as GR and GPx, the direct reactivity of DCI-PS towards purified TrxR could not be explored due to the non-availability of the pure protein from commercial suppliers at reasonable/affordable prices.…”

“…1b). 30 Interestingly, the observations from all the experimental and theoretical studies collectively provide a sufficient body of evidence for the predominant contribution/participation of TrxR for the turn-on fluorogenic processes for the sustained release of H 2 S with concomitant release of NIR fluorophore. The involvement of TrxR is not only beneficial for the sustained donation of H 2 S for the H 2 S-mediated therapeutics in disease models, but the strategy can also be useful for estimating the relative cellular expression level of TrxR in different organspecific cancers as well as inflammatory diseases with the highly tissue-penetrating NIR dye.…”

“…1b). 30 These probes can effectively estimate the overexpression level of TrxR in disease conditions such as cancer. Similarly, the turn-on fluorogenic probes for other antioxidant enzymes such as GR and GPx were also developed recently for estimating their cellular expression levels.…”

“…The choice of DTT, a small-molecule dithiol as a functional mimic of TrxR was based on the structural and nucleophilic functional similarities. 30,35 As the reduction of selenenyl sulfide (-Se-S-) bond in TrxR generates the highly nucleophilic and adjacent selenolate-thiol moiety, the reduced DTT also exists as highly nucleophilic and adjacent dithiol moieties (Fig. 2).…”

Discussions of Fluorescence in Selenium Chemistry: Recently Reported Probes, Particles, and a Clearer Biological Knowledge

“…In this case, of importance for medicinal studies is that the binding affinity was measured as 15.9 µM. Imaging using in vitro biological results supported the superiority in analyte detection of the Se-Se-linked derivative over that of the S-S-linked derivative [49]. The report by Mafireyi et al discusses a probe in which the analyte is, in fact, a full protein.…”