Three-dimensional structure of a mammalian thioredoxin reductase: Implications for mechanism and evolution of a selenocysteine-dependent enzyme

Thioredoxin reductases (TrxRs) from mammalian cells contain an essential se lenocysteine residue in the conserved C-terminal sequence Gly-Cys-SeCys-Gly forming a selenenylsulfide in the oxidized enzyme. Reduction by NADPH gene rates a selenolthiol, which is the active site in reduction of Trx. The thr ee-dimensional structure of the SeCys498Cys mutant of rat TrxR in complex w ith NADP(+) has been determined to 3.0-Angstrom resolution by x-ray crystal lography. The overall structure is similar to that of glutathione reductase (GR), including conserved amino acid residues binding the cofactors FAD an d NADPH. Surprisingly, all residues directly interacting with the substrate glutathione disulfide in GR are conserved despite the failure of glutathio ne disulfide to act as a substrate for TrxR. The 16-residue C-terminal tail , which is unique to mammalian TrxR, folds in such a way that it can approa ch the active site disulfide of the other subunit in the dimer. A model of the complex of TrxR with Trx suggests that electron transfer from NADPH to the disulfide of the substrate is possible without large conformational cha nges. The C-terminal extension typical of mammalian TrxRs has two functions : (i) it extends the electron transport chain from the catalytic disulfide to the enzyme surface, where it can react with Trx, and (ii) it prevents th e enzyme from acting as a GR by blocking the redox-active disulfide. Our re sults suggest that mammalian TrxR evolved from the GR scaffold rather than from its prokaryotic counterpart. This evolutionary switch renders cell gro wth dependent on selenium.