4UP3

Crystal structure of the mutant C140S,C286Q thioredoxin reductase from Entamoeba histolytica

Summary for 4UP3

• Entry DOI: 10.2210/pdb4up3/pdb
• Descriptor: THIOREDOXIN REDUCTASE, FLAVIN-ADENINE DINUCLEOTIDE, NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE, ... (5 entities in total)
• Functional Keywords: oxidoreductase, redox metabolism, oxidative stress
• Biological source: ENTAMOEBA HISTOLYTICA
• Total number of polymer chains: 2
• Total formula weight: 70701.06

Authors

Podust, L.M.,Vieira, D.F. (deposition date: 2014-06-11, release date: 2015-07-01, Last modification date: 2024-01-10)

Primary citation

Parsonage, D.,Sheng, F.,Hirata, K.,Debnath, A.,Mckerrow, J.H.,Reed, S.L.,Abagyan, R.,Poole, L.B.,Podust, L.M.
X-Ray Structures of Thioredoxin and Thioredoxin Reductase from Entamoeba Histolytica and Prevailing Hypothesis of the Mechanism of Auranofin Action.
J.Struct.Biol., 194:180-, 2016

PubMed Abstract: The anti-arthritic gold-containing drug Auranofin is lethal to the protozoan intestinal parasite Entamoeba histolytica, the causative agent of human amebiasis, in both culture and animal models of the disease. A putative mechanism of Auranofin action proposes that monovalent gold, Au(I), released from the drug, can bind to the redox-active dithiol group of thioredoxin reductase (TrxR). Au(I) binding in the active site is expected to prevent electron transfer to the downstream substrate thioredoxin (Trx), thus interfering with redox homeostasis in the parasite. To clarify the molecular mechanism of Auranofin action in more detail, we determined a series of atomic resolution X-ray structures for E. histolytica thioredoxin (EhTrx) and thioredoxin reductase (EhTrxR), the latter with and without Auranofin. Only the disulfide-bonded form of the active site dithiol (Cys(140)-Cys(143)) was invariably observed in crystals of EhTrxR in spite of the addition of reductants in various crystallization trials, and no gold was found associated with these cysteines. Non-catalytic Cys(286) was identified as the only site of modification, but further mutagenesis studies using the C286Q mutant demonstrated that this site was not responsible for inhibition of EhTrxR by Auranofin. Interestingly, we obtained both of the catalytically-relevant conformations of this bacterial-like, low molecular weight TrxR in crystals without requiring an engineered disulfide linkage between Cys mutants of TrxR and Trx (as was originally done with Escherichia coli TrxR and Trx). We note that the -CXXC- catalytic motif, even if reduced, would likely not provide space sufficient to bind Au(I) by both cysteines of the dithiol group.

PubMed: 26876147
DOI: 10.1016/J.JSB.2016.02.015

Experimental method

X-RAY DIFFRACTION (1.44 Å)