Asn112 in Plasmodium falciparum glutathione S-transferase is essential for induced reversible tetramerization by phosphate or pyrophosphate |
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Authors: | Indalecio Quesada-Soriano,Carmen Baró nRamiro Té llez-Sanz,Federico Garcí a-MarotoLuis Garcí a-Fuentes |
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Affiliation: | Department of Chemistry and Physics, University of Almería, Research Center for Agrifood Biotechnology (BITAL), Agrifood Campus of International Excellence ceiA3, Ctra de Sacramento s/n, 04120 Almería, Spain |
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Abstract: | The glutathione S-transferase from Plasmodium falciparum presents distinct features which are absent from mammalian GST isoenzyme counterparts. Most apparent among these are the ability to tetramerize and the presence of a flexible loop. The loop, situated between the 113–119 residues, has been reported necessary for the tetramerization process. In this article, we report that a residue outside of this loop, Asn112, is a key to the process — to the point where the single Asn112Leu mutation prevents tetramerization altogether. We propose that a structural pattern involving the interaction of the Asn112 and Lys117 residues from two neighboring subunits plays a role in keeping the tetramer structure stable. We also report that, for the tetramerization of the wild-type PfGST to occur, phosphate or pyrophosphate anions must be present. In other words, tetramerization is a phosphate- or pyrophosphate-induced process. Furthermore, the presence of magnesium reinforces this induction. We present experimental evidence for these claims as well as a preliminary calorimetric and kinetic study of the dimeric Asn112Leu PfGST mutant. We also propose a putative binding site for phosphate or pyrophosphate anions through a comparative structural analysis of PfGST and pyrophosphatases from several organisms. Our results highlight the differences between PfGST and the human isoenzymes, which make the parasite enzyme a suitable antimalarial target. |
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Keywords: | GST, glutathione S-transferase PfGST, glutathione S-transferase from Plasmodium falciparum ITC, isothermal titration calorimetry GSH, reduced glutathione CDNB, 1-chloro-2,4-dinitrobenzene SEC, size-exclusion chromatography DSC, differential scanning calorimetry |
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