Inhibition of Schistosoma mansoni Thioredoxin-glutathione Reductase by Auranofin

Abstract

Schistosomiasis is a parasitic disease affecting over 200 million people currently treated with one drug, praziquantel. A possible drug target is the seleno-protein thioredoxin-glutathione reductase (TGR), a key enzyme in the pathway of the parasite for detoxification of reactive oxygen species. The enzyme is a unique fusion of a glutaredoxin domain with a thioredoxin reductase domain, which contains a selenocysteine (Sec) as the penultimate amino acid. Auranofin (AF), a gold-containing compound already in clinical use as an anti-arthritic drug, has been shown to inhibit TGR and to substantially reduce worm burden in mice. Using x-ray crystallography we solved (at 2.5 Å resolution) the structure of wild type TGR incubated with AF. The electron density maps show that the actual inhibitor is gold, released from AF. Gold is bound at three different sites not directly involving the C-terminal Sec residue; however, because the C terminus in the electron density maps is disordered, we cannot exclude the possibility that gold may also bind to Sec. To investigate the possible role of Sec in the inactivation kinetics, we tested the effect of AF on a model enzyme of the same superfamily, i.e. the naturally Sec-lacking glutathione reductase, and on truncated TGR. We demonstrate that the role of selenium in the onset of inhibition by AF is catalytic and can be mimicked by an external source of selenium (benzeneselenol). Therefore, we propose that Sec mediates the transfer of gold from its ligands in AF to the redox-active Cys couples of TGR.

Mapping the Catalytic Cycle of Schistosoma mansoni Thioredoxin Glutathione Reductase by X-ray Crystallography

“…Until now, we crystallized three different variants of SmTGR in four different space groups (this paper and Refs. 13,17). 5 The data show different crystallographic contacts but always the same overall architecture, suggesting the structure in solution and in the crystal is the same.…”

“…The schistosomal antioxidant system lacks the typical mammalian TR and GR enzymes, which are both replaced by SmTGR; this peculiarity revealed that SmTGR may be a target of choice for the development of new antischistosomal drugs (5,13). Moreover, TGR enzymes with similar roles have been isolated in other human parasites, such as Taenia (14) and Echinococcus (15).…”

The C Terminus of the Catalytic Domain of Type A Botulinum Neurotoxin May Facilitate Product Release from the Active Site

“…We therefore performed some analysis of the full-length LcB structure, 437 residues of which could be accounted for in the electron density (Protein Data Bank code 2ETF). 5 The core structures of LcA and LcB are superimposable as are all serotypes with very little C␣ root mean square deviation. The presence of the loop between residues 415 and 425 makes the long helix (residues 425-437) very floppy so it can assume any orientation in LcB.…”

“…The LcB structure is from the Protein Data Bank (code 2ETF). 5 Green, C-terminal helix (residues 425-437); red, C-terminal loop; yellow, 60, 200, and 370 loops as indicated; blue, zinc atom bound at the active site.…”

“…Recent years have witnessed a tremendous proliferation of three-dimensional structure determinations by the advent of high throughput x-ray crystallography soon after the sequence and adequate expression of a protein became available (1,2). In some proteins, no electron density can be observed for stretches of the amino acid sequence especially at the N or C terminus (3)(4)(5)(6)(7)(8). Thus, a functional role for such regions is not always discernible from their three-dimensional structures.…”