Regulation of Thioredoxin Peroxidase Activity by C-terminal Truncation

Thioredoxin peroxidase is a member of peroxiredoxin (Prx) family, which uses a thioredoxin (Trx) as an immediate electron donor for the reduction of peroxide. We have identified C-terminal truncated TPx from Schizosaccharomyces pombe and also have found the truncated form is significantly tenacious against the inactivation of H2O2 than the intact form. Peroxidase assay of a series of recombinant C-terminal truncation mutants (Δ192, Δ191, Δ188, Δ184, Δ176, and Δ165) revealed that TPx could be inactivated (Δ192), reactivated (Δ191–Δ176) and reinactivated (Δ165) by serial truncation from C-terminus. We did not find any significant kinetic difference among reactivated forms; however, distinctive loss of affinity to H2O2 (Km = 5 μM) than that of the intact form (<<5 μM, undeterminable) was monitored. Characterization of a series of Lys191 point mutants manifested that the loss of affinity caused by a deprivation of positive charge born in Lys191 and the loss of affinity resulted in the resistibility to H2O2. Disk inhibition assay with S. pombe cells overexpressing wild-type, Δ192 and Δ191 mutants evidenced that the truncated forms functioning in vitro as well as in vivo.