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Thioredoxin reductase in redox regulation and genetic code

Anton A Turanov, University of Nebraska - Lincoln

Abstract

Thioredoxin reductase (TR) and thioredoxin (Trx) define a major cellular redox system that maintains intracellular redox balance. Both cytosolic (TR1, Trx1) and mitochondrial (TR3, Trx2) enzymes are essential in mammals.^ We identified and characterized in detail the subcellular localization of three TR3 forms that are generated by alternative first exon splicing and differ in their N-terminal sequences. Recombinant selenoprotein forms of TR1 and TR3 were generated and regulation of their activities by metal ions investigated. Both enzymes were inhibited by zinc, but not by calcium or cobalt. In addition, we developed a proteomic method to identify targets of mammalian TRs. Trx1 was the major target for both TR1 and TR3 in rat and mouse liver cytosolic fractions and could be specifically enriched from tissue extracts and body liquids on TR affinity columns. To examine the roles of TRs in redox regulation, knockout mice in which selenocysteine (Sec) tRNA gene was disrupted specifically in the liver were employed. We found that the levels of TR1 and TR3 were reduced in the knockout samples, whereas the levels of Trx1 and Trx2 were elevated; however, Trxs were in the oxidized state in the knockout samples. Overall, these data provided evidence for key roles of TRs in redox homeostasis. ^ In ciliated protozoa Euplotes crassus, cysteine (Cys) is encoded by three codons, UGA, UGU and UGC. UGA is a stop signal in the universal genetic code, and this codon can also code for the 21st amino acid, Sec, in all three domains of life. We found that in Euplotes crassus the UGA codon specifies the insertion of both Cys and Sec. We characterized the E. crassus thioredoxin reductase gene, in which the first six UGA codons code for Cys and the seventh for Sec. Our data show that E. crassus utilizes UGA for insertion of both Cys and Sec, establishing it as the first known organism that utilizes one codon to code unambiguously for two amino acids.^

Subject Area

Chemistry, Biochemistry

Recommended Citation

Turanov, Anton A, "Thioredoxin reductase in redox regulation and genetic code" (2008). ETD collection for University of Nebraska - Lincoln. AAI3336556.
http://digitalcommons.unl.edu/dissertations/AAI3336556

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