Abstract
Schistosomiasis is the second most widespread human parasitic disease. It is principally treated with one drug, praziquantel, that is administered to 100 million people each year; less sensitive strains of schistosomes are emerging. One of the most appealing drug targets against schistosomiasis is thioredoxin glutathione reductase (TGR). This natural chimeric enzyme is a peculiar fusion of a glutaredoxin domain with a thioredoxin selenocysteine (U)-containing reductase domain. Selenocysteine is located on a flexible C-terminal arm that is usually disordered in the available structures of the protein and is essential for the full catalytic activity of TGR. In this study, we dissect the catalytic cycle of Schistosoma mansoni TGR by structural and functional analysis of the U597C mutant. The crystallographic data presented herein include the following: the oxidized form (at 1.9 Å resolution); the NADPH- and GSH-bound forms (2.3 and 1.9 Å, respectively); and a different crystal form of the (partially) reduced enzyme (3.1 Å), showing the physiological dimer and the entire C terminus of one subunit. Whenever possible, we determined the rate constants for the interconversion between the different oxidation states of TGR by kinetic methods. By combining the crystallographic analysis with computer modeling, we were able to throw further light on the mechanism of action of S. mansoni TGR. In particular, we hereby propose the putative functionally relevant conformational change of the C terminus after the transfer of reducing equivalents from NADPH to the redox sites of the enzyme.
Highlights
Schistosomes are human platyhelminth parasites causing Schistosomiasis, a severe disease still classified among the major causes of mortality in tropical and subtropical countries, affecting more than 200 million people [1]
The main evidence suggesting that targeting the thiol redox pathway of the parasite may represent a profitable starting point for rational drug design is as follows: (i) schistosomes, living in the human bloodstream, are subjected to endogenous reactive oxygen species but are exposed to radicals produced by the host immune response; and (ii) the thiol redox pathway employed by the worms for the enzymatic reduction of the reactive oxygen species is different from its human counterpart [5, 6]
Concluding Remarks—The data reported in this study describe the complete catalytic cycle of SmTGR, a complex enzyme, based on structural data and modeling
Summary
The two domains work either coupled or independently to catalyze the reduction of a large number of oxidized substrates. The C-terminal tail of each subunit, which is close to the FAD active site of the adjacent subunit, contains the Sec residue [17]. The catalytic mechanism of TGR has not been fully elucidated, an hypothesis has been published for the mouse enzyme [18]
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