title: Thioredoxin-mimetic peptides as catalysts of S-denitrosylation and anti-nitrosative stress agents.
abstract: S-nitrosylation, the coupling of a nitric oxide moiety to a reactive cysteine residue to form an S-nitrosothiol (SNO), is an important posttranslational mechanism for regulating protein activity. Growing evidence indicates that hyper-S-nitrosylation may contribute to cellular dysfunction associated with various human diseases. It is also increasingly appreciated that thioredoxin and thioredoxin reductase play significant roles in the cellular catabolism of SNO and protection from nitrosative stress. Here, we investigated the SNO reductase activity and protective effects of thioredoxin-mimetic peptides (TXMs), Ac-Cys-Pro-Cys-amide (CB3) and Ac-Cys-Gly-Pro-Cys-amide (CB4), both under cell-free conditions and in nitrosatively stressed cultured cells. In vitro biochemical analyses revealed that the TXM peptides reduced small-molecule SNO compounds, such as S-nitrosoglutathione (GSNO), and acted as general and efficient protein-denitrosylating agents. In particular, CB3 was found to be a highly potent SNO-metabolizing agent. Notably, CB3 mimicked the activity of thioredoxin by coupling with thioredoxin reductase to enhance GSNO reduction. Moreover, in a cell-free lysate system, both CB3 and CB4 synergized with an NADPH-dependent activity to denitrosylate proteins. Further investigation revealed that the TXM peptides protect the peroxiredoxin-thioredoxin system from SNO-dependent inhibition. Indeed, SNO-inhibited Prx1 was efficiently denitrosylated and reactivated by CB3 or CB4. In addition, CB3 protected thioredoxin reductase from SNO-mediated inactivation both in vitro and in intact cells. Finally, CB3 and CB4 partially rescued human neuroblastoma SH-SY5Y cells and rat insulinoma INS-1 832/13 cells from GSNO-induced growth inhibition. Collectively, the present findings indicate the efficient denitrosylation activity and protective effects of TXM peptides and suggest their potential therapeutic value in treating pathological conditions related to nitrosative stress.
authors.affiliation: Department of Biochemistry, Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa 31096, Israel.
authors.forename: Gali
authors.initials: G
authors.lastname: Kronenfeld
authors.affiliation: Department of Biochemistry, Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa 31096, Israel.
authors.forename: Rotem
authors.initials: R
authors.lastname: Engelman
authors.affiliation: Department of Biochemistry, Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa 31096, Israel.
authors.forename: Pnina
authors.initials: P
authors.lastname: Weisman-Shomer
authors.affiliation: Department of Biological Chemistry, Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel. Electronic address: daphne.atlas@mail.huji.ac.il.
authors.forename: Daphne
authors.initials: D
authors.lastname: Atlas
authors.affiliation: Department of Biochemistry, Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa 31096, Israel. Electronic address: benhar@tx.technion.ac.il.
authors.forename: Moran
authors.initials: M
authors.lastname: Benhar
mesh_terms: D002384:Catalysis; D002460:Cell Line; D006801:Humans; D018716:Molecular Mimicry; D008970:Molecular Weight; D015538:Nitrosation; D013879:Thioredoxins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D013879:Thioredoxins
keywords: Denitrosylation; Free radicals; Nitrosative stress; Thiols; Thioredoxin mimetic peptides