Thiol-targeted introduction of selenocysteine to polypeptides for synthesis of glutathione peroxidase mimics

Because the seleno-l-cysteine (SeCys or Sec) insertion into selenoproteins occurs by a specific translational control process, it is quite difficult to express the SeCys-containing polypeptides even by the state-of-the-art genetic engineering techniques. In this paper, we describe a convenient synthetic method for the selective introduction of a SeCys derivative to polypeptides under physiological conditions. One SeCys residue in the seleno-l-cystine (SeCys-Se-Se-SeCys) methyl ester was first substituted with the Boc-protected penicillamine (Pen) methyl ester to form selenenylsulfide (SeCys-Se-S-Pen), an intermediate in the cellular glutathione peroxidase (GPx) catalytic cycle. Subsequently, the SeCys-Pen was coupled with the thiol-specific N-carboxymethylmaleimide through the α-amino group of the SeCys {[2-(N-maleimidyl)-1-oxo-ethyl-SeCys-methyl-Se-yl]-S-Pen methyl ester, MOE-SeCys-Pen}. The use of the MOE-SeCys-Pen allowed the selective introduction of the SeCys moiety to human serum albumin by alkylation of the thiol at its cysteine34, which generated the GPx-like activity responsible for the selenium atom in the MOE-SeCys-Pen. Consequently, this synthetic method will allow generating SeCys-containing artificial polypeptides with a GPx-like activity.