Regulation of the extracellular antioxidant selenoprotein plasma glutathione peroxidase (GPx-3) in mammalian cells.
Plasma glutathione peroxidase (GPx-3) is a selenocysteine-containing extracellular antioxidant protein that catalyzes the reduction of hydrogen peroxide and lipid hydroperoxides. Selenoprotein expression involves the alternate recognition of a UGA codon as a selenocysteine codon and requires signals in the 3'-untranslated region (UTR), including a selenocysteine insertion sequence (SECIS), as well as specific translational cofactors. To ascertain regulatory determinants of GPx-3 expression and function, we generated recombinant GPx-3 (rGPX-3) constructs with various 3'-UTR, as well as a Sec73Cys mutant. In transfected Cos7 cells, the Sec73Cys mutant was expressed at higher levels than the wild type rGPx-3, although the wild type rGPx-3 had higher specific activity, similar to plasma purified GPx-3. A 3'-UTR with only the SECIS was insufficient for wild type rGPx-3 protein expression. Selenocompound supplementation and co-transfection with SECIS binding protein 2 increased wild type rGPx-3 expression. These results demonstrate the importance of translational mechanisms in GPx-3 expression.
Ottaviano
Filomena G
FG
Whitaker Cardiovascular Institute and Evans Department of Medicine, Boston University School of Medicine, Boston, MA, USA.
Tang
Shiow-Shih
SS
Handy
Diane E
DE
Loscalzo
Joseph
J
D020413:3' Untranslated Regions / Q000378:metabolism; D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D005979:Glutathione Peroxidase / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012643:Selenium / Q000494:pharmacology; D051140:Selenoproteins / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D020413:3' Untranslated Regions; D000975:Antioxidants; D011994:Recombinant Proteins; D051140:Selenoproteins; C486484:GPX3 protein, human; D005979:Glutathione Peroxidase; D012643:Selenium