Oxidized Forms of Ergothioneine Are Substrates for Mammalian Thioredoxin Reductase

Abstract

Ergothioneine (EGT) is a sulfur-containing amino acid analog that is biosynthesized in fungi and bacteria, accumulated in plants, and ingested by humans where it is concentrated in tissues under oxidative stress. While the physiological function of EGT is not yet fully understood, EGT is a potent antioxidant in vitro. Here we report that oxidized forms of EGT, EGT-disulfide (ESSE) and 5-oxo-EGT, can be reduced by the selenoenzyme mammalian thioredoxin reductase (Sec-TrxR). ESSE and 5-oxo-EGT are formed upon reaction with biologically relevant reactive oxygen species. We found that glutathione reductase (GR) can reduce ESSE, but only with the aid of glutathione (GSH). The reduction of ESSE by TrxR was found to be selenium dependent, with non-selenium-containing TrxR enzymes having little or no ability to reduce ESSE. In comparing the reduction of ESSE by Sec-TrxR in the presence of thioredoxin to that of GR/GSH, we find that the glutathione system is 10-fold more efficient, but Sec-TrxR has the advantage of being able to reduce both ESSE and 5-oxo-EGT directly. This represents the first discovered direct enzymatic recycling system for oxidized forms of EGT. Based on our in vitro results, the thioredoxin system may be important for EGT redox biology and requires further in vivo investigation.

Computational Insights into the Regeneration of Ovothiol and Ergothioneine and Their Selenium Analogues by Glutathione

“…These results are in agreement with the past experimental work that has shown that OSH acts in tandem with glutathione, where OSH neutralizes oxidants forming OSSO with GSH then used to maintain ovothiol in the reduced state . Moreover, previous experimental work has shown that glutathione reductase can reduce ESSE, but only with the aid of glutathione . However, looking at Figures and , the second disulfide exchange reaction is generally slower than the respective first disulfide exchange reaction.…”

“… 77 Moreover, previous experimental work has shown that glutathione reductase can reduce ESSE, but only with the aid of glutathione. 78 However, looking at Figures 2 and 5 , the second disulfide exchange reaction is generally slower than the respective first disulfide exchange reaction. Specifically, comparing Reactions 10 and 2, Reactions 10 and 3, Reactions 12 and 6, and Reactions 12 and 7, the Gibbs activation energies of the second step are on average 8.2 kJ mol –1 more endergonic.…”

Discovery and structure of a widespread bacterial ABC transporter specific for ergothioneine

“…This finding is consistent with the very weak, but measurable (K d ≈ mM) binding of EgtUC to GB and Lhercynine observed here. On the other hand, we have not yet elucidated the potential impact of oxidation or methylation of the ET thione S to create sulfonylated ET, or S-methylated ET, respectively 67 , or insertion of an oxygen atom to the C5-position to create 5-oxo-ET 9 , or Se substitution of the S atom in ET in selenoneine 68,69 , on EgtUC ligand binding affinity. Indeed, the ligand specificity of EgtU homologs in other SSN cluster 2 SBDs remains to be experimentally validated 57 .…”

“…1). This thiol-thione tautomerization significantly increases the thiol-disulfide reduction potential of ET relative to other LMW thiols thus endowing ET with properties of a highly effective scavenger of myriad ROS 8,9 . ET also chelates transition metals including Cu + , Cu 2+ , and Fe 2+ and thus may impact metal and metal oxidation state speciation in cells [10][11][12] .ET biosynthesis has been reported to occur only in select filamentous fungi including Neurospora crassa 13 , certain cyanobacteria 14 ,…”

Ergothioneine: an underrecognised dietary micronutrient required for healthy ageing?

“…Ergothioneine deactivates singlet-oxygen species at higher rates than other thiols (49) , and it has been hypothesised that the primary function of ergothioneine may be to prevent damage at intracellular sites of high singlet-oxygen generation (26) . Redox repair of the oxidised forms (ergothioneine disulfide and 5-oxo-ergothioneine) of ergothioneine can be rapidly provided by ascorbate (48) or can be achieved enzymatically by either glutathione reductase in presence of glutathione, or the selenoenzyme thioredoxin reductase (50) .…”