Can Selenoenzymes Resist Electrophilic Modification? Evidence from Thioredoxin Reductase and a Mutant Containing α-Methylselenocysteine

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Chemistry and Biochemistry


Mathematics and Natural Sciences


Selenocysteine (Sec) is the 21st proteogenic amino acid in the genetic code. Incorporation of Sec into proteins is a complex and bioenergetically costly process that evokes the question: “Why did nature choose selenium?” An answer that has emerged over the past decade is that Sec confers resistance to irreversible oxidative inactivation by reactive oxygen species (ROS). Here, we explore the question of whether this concept can be broadened to include resistance to reactive electrophilic species (RES) since oxygen and related compounds are merely a subset of RES. To test this hypothesis we inactivated mammalian thioredoxin reductase (Sec-TrxR), a mutant containing alpha-methylselenocysteine ((αMe)Sec-TrxR), and a cysteine-ortholog TrxR (Cys-TrxR) with various electrophiles including acrolein, 4-hydroxynonenal, and curcumin. Our results show that the acrolein-inactivated Sec-TrxR and the (αMe)Sec-TrxR mutant could regain 25% and 30% activity respectively, when incubated with 2 mM H2O2 and 5 mM imidazole. In contrast, the Cys-TrxR did not regain activity under the same conditions. We posit that Sec-enzymes can undergo a repair process via β-syn selenoxide elimination that ejects the electrophile, leaving the enzyme in the oxidized selenosulfide state. (αMe)Sec-TrxR was created by incorporating the non-natural amino acid ((αMe)Sec into TrxR by semisynthesis and allowed for rigorous testing of our hypothesis. This Sec-derivative enables higher resistance to both oxidative and electrophilic inactivation because it lacks a backbone Cα-H, which prevents loss of selenium through formation of dehydroalanine. This is the first time this unique amino acid has been incorporated into an enzyme and is example of state-of-the-art protein engineering.

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