Characterization of a mammalian peroxiredoxin that contains one conserved cysteine

A new type of peroxidase enzyme, named thioredoxin peroxidase (TPx), that reduces H₂O₂ with the use of electrons from thioredoxin and contains two essential cysteines was recently identified. TPx homologs, termed peroxiredoxin (Prx), have also been identified and include several proteins, designated 1-Cys Prx, that contain only one conserved cysteine. Recombinant human 1-Cys Prx expressed in and purified from Escherichia coli has now been shown to reduce H₂O₂ with electrons provided by dithiothreitol. Furthermore, human 1-Cys Prx transiently expressed in NIH 3T3 cells was able to remove intracellular H₂O₂ generated in response either to the addition of exogenous H₂O₂ or to treatment with platelet-derived growth factor. The conserved Cys⁴⁷-SH group was shown to be the site of oxidation by H₂O². Thus, mutation of Cys⁴⁷ to serine abolished peroxidase activity. Moreover, the oxidized intermediate appears to be Cys-SOH. In contrast to TPx, in which one of the two conserved cysteines is oxidized to Cys-SOH and then immediately reacts with the second conserved cysteine of the second subunit of the enzyme homodimer to form an intermolecular disulfide, the Cys-SOH of 1-Cys Prx does not form a disulfide. Neither thioredoxin, which reduces the disulfide of TPx, nor glutathione, which reduces the Cys-SeOH of oxidized glutathione peroxidase, was able to reduce the Cys-SOH of l-Cys Prx and consequently could not support peroxidase activity. Human 1-Cys Prx was previously shown to exhibit a low level of phospholipase A₂ activity at an acidic pH; the enzyme was thus proposed to be lysosomal, and Ser³² was proposed to be critical for lipase function. However, the mutation of Set³² or Cys⁴⁷ has now been shown to have no effect on the lipase activity of 1- Cys Prx, which was also shown to be a cytosolic protein. Thus, the primary cellular function of 1-Cys Prx appears to be to reduce peroxides with the use of electrons provided by an as yet unidentified source; the enzyme therefore represents a new type of peroxidase.