Ebselen: A substrate for human thioredoxin reductase strongly stimulating its hydroperoxide reductase activity and a superfast thioredoxin oxidant

Abstract

Ebselen [2-phenyl-1,2-benzisoselenazol-3(2 H )-one], a seleno-organic compound with glutathione peroxidase-like activity is used in clinical trials against stroke. Human and bovine TrxR catalyzed the reduction of ebselen to ebselen selenol by NADPH with an apparent K M -value of 2.5 μM and a k cat of 588 min −1 . The addition of thioredoxin (Trx) stimulated the TrxR-catalyzed reduction of ebselen several-fold. This result was caused by a very fast oxidation of reduced Trx by ebselen with a rate constant in excess of 2 × 10 7 M −1 s −1 . This rate is orders of magnitude faster than the reaction of dithiol Trx with insulin disulfides. Ebselen competed with disulfide substrates for reduction by Trx and, therefore, acted as an inhibitor of protein disulfide reduction by the Trx system. The inherent H 2 O 2 reductase activity of mammalian TrxR dependent on its active-site selenocysteine residue was stimulated 10-fold by 2 μM ebselen and 25-fold in the additional presence of 5 μM Trx. Furthermore, the apparent K M -value of TrxR for H 2 O 2 was lowered 25-fold to about 100 μM. Our results demonstrate that ebselen is a TrxR peroxidase which, in the presence of Trx, acted as a mimic of a peroxiredoxin. The activity with TrxR and oxidation of reduced Trx offer mechanistic explanations for the in vivo effects of ebselen as an antioxidant and anti-inflammatory agent. Our results demonstrate that the mechanism of action of ebselen may be predominantly via the Trx system rather than via glutathione.