Inhibition of thioredoxin and thioredoxin reductase by 4-hydroxy-2-nonenal in vitro and in vivo.
Lipid peroxidation is a cellular process that takes place under physiological conditions and particularly after oxidative stress. 4-Hydroxy-2-nonenal (HNE), a major end product of lipid peroxidation, is known to exert a multitude of biological effects and has high reactivity to various cellular components, including DNA and protein. The thioredoxin system, composed of the selenoenzyme thioredoxin reductase (TrxR), thioredoxin (Trx), and NADPH, plays a key role in redox regulation and is involved in many signaling pathways. The selenocysteine (Sec) and cysteine (Cys) residues (Cys-496/Sec-497) in the active site of TrxR and a pair of Cys residues (Cys-32/Cys-35) in Trx are sensitive to various alkylating reagents. Herein, we report a mechanistic study on the inhibition of rat TrxR by HNE. The inhibition occurs with TrxR only in its reduced form and persists after removal of HNE. Inhibition of TrxR by HNE added to cultured HeLa cells is also observed. In addition, HNE inactivates reduced Escherichia coli Trx irreversibly. We proved that the redox residues (Cys-496/Sec-497 in TrxR and Cys-32/Cys-35 in Trx) were primary targets for HNE modification. The covalent adducts formed between HNE and Trx were also confirmed by mass spectrum. Because the thioredoxin system is one of the core regulation enzymes of cells' function, inhibition of both TrxR and Trx by HNE provides a possibly novel mechanism for explanation of its cytotoxic effect and signaling activity, as well as the further damage indirectly caused under oxidative stress conditions.
Fang
Jianguo
J
Medical Nobel Institute for Biochemistry, Department of Medical Biochemistry and Biophysics, Karolinska Institute, SE-17177 Stockholm, Sweden. jianguo.fang@mbb.ki.se
Holmgren
Arne
A
D000447:Aldehydes / Q000494:pharmacology*; D001665:Binding Sites; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D006367:HeLa Cells; D006801:Humans; D007328:Insulin / Q000378:metabolism; D007700:Kinetics; D010084:Oxidation-Reduction; D017279:Selenocysteine / Q000737:chemistry / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013880:Thioredoxin-Disulfide Reductase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D013879:Thioredoxins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism
D000447:Aldehydes; D007328:Insulin; D017279:Selenocysteine; D013879:Thioredoxins; D013880:Thioredoxin-Disulfide Reductase; C027576:4-hydroxy-2-nonenal; D003545:Cysteine