Abstract
Aims: Thioredoxin reductase 1 (TrxR1) is a cancer target and essential selenoprotein that defends the cell against reactive oxygen species and regulates cellular signaling and redox pathways. Previous cell-based studies correlated TrxR1 acetylation with modulated cellular reduction activity, yet the function of specific acetylation sites on TrxR1 remains unknown.Innovation: We produced site-specifically acetylated TrxR1 variants that also contain selenocysteine (Sec). We demonstrated efficient high-fidelity protein synthesis with 22 different amino acids by simultaneous UAG codon reassignment to Nɛ-acetyl-lysine and UGA codon recoding to Sec.Results: We characterized TrxR1 variants acetylated at physiologically relevant sites and found that single acetylation sites increased TrxR1 activity, enhancing the apparent catalytic rate up to 2.7-fold. The activity increase in acetylated TrxR1 (acTrxR1) is reversible and is reduced following deacetylation with histone deacetylase.Conclusion: Here we present a novel mechanism through which acetylation increases TrxR1 activity by destabilizing low-activity TrxR1 multimers, increasing the population of active dimeric TrxR1. Antioxid. Redox Signal. 29, 377–388.
Highlights
Human cells naturally defend against reactive oxygen species (ROS) using a network of redox enzymes, including the selenoprotein thioredoxin reductase 1 (TrxR1) [38]
The Sec synthesis and insertion system is endogenous to Escherichia coli, encoded in the E. coli genome (SelA, SelB, SelC, SelD, SerRS). 3¢ UTR, 3¢ untranslated region; acK, Ne-acetyl-lysine; acKRS, mutant pyrrolysyl-tRNA synthetase (Ne-acetyl-lysine-tRNA synthetase); Sec, selenocysteine; SecIS, selenocysteine insertion sequence; Thioredoxin reductase 1 (TrxR1), thioredoxin reductase 1
Lysine acetylation is found in all domains of life [10, 40, 51] with 36,000 acetylation sites identified in human, rat, and mouse cells [7, 21]
Summary
Human cells naturally defend against reactive oxygen species (ROS) using a network of redox enzymes, including the selenoprotein thioredoxin reductase 1 (TrxR1) [38]. Selenoproteins contain the 21st genetically encoded amino acid, selenocysteine (Sec). The Sec550 residue in TrxR1 is required for most of its activity [42] in reducing oxidized compounds, for example, ubiquinone [53], and oxidatively damaged cellular proteins via a redox-coupled reaction with thioredoxin (Trx). TrxR1 catalyzes the transfer of electrons from b-nicotinamide adenine dinucleotide 2¢-phosphate (NADPH) to Trx. The reduced Trx in turn resolves oxidized species and reduces cellular proteins. The resulting oxidized Trx is recycled by the TrxR1 enzyme. In addition to ROS defense, the Trx system is involved in regulating gene expression, embryonic development, cell proliferation, and apoptosis [27]
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