Title:
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Trypanosomatid chemotherapy : crystal structure-based drug design of phosphoglycerate mutase from Leishmania mexicana and Trypanosoma brucei
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2,3-bisphosphoglycerate-independent phosphoglycerate mutase from Leishmania mexicana (Lm iPGAM) is a member of the divalent metal-dependent phosphatase superfamily, and catalyses the interconversion of 3- and 2-phosphoglycerates in the glycolytic pathway via a phosphoserine intermediate. The determination of the structure of Lm iPGAM provides a foundation for structure-based drug design studies that are currently in progress to develop novel drugs to tackle diseases caused by L. mexicana and Trypanosoma brucei parasites. Recent RNAi experiments have shown that iPGAM is essential for the survival of bloodstream-form T. brucei, L. mexicana and T. brucei iPGAMs share 74% sequence identity, but have no common features with cofactor-dependent PGAM from humans. L. mexicana iPGAM is one of the rare cobalt enzymes that have been well characterised crystallographically. Comparison with the crystal structure of manganese-dependent Bacillus stearothermophilus iPGAM shows that the active site architectures of both enzymes are very similar, although the sequence identity is only 33%. Sequence comparisons give clues about metal specificity, especially by the presence of an insertion at Tyr210 in Lm iPGAM that cause His360 to adapt a position where it can form a H-bond with the phosphor group of the substrate/product. Inspection of these structures shows that the metal specificity of each enzyme depends on the active site volume, which corresponds to the chemical nature of a particular metal. The metal-free Lm iPGAM can be reactivated fully only by cobalt, but not by manganese or zinc. These metals cause the metal coordination geometries to change, as revealed crystallographically, and thus may cause the loss of enzyme activity. Moreover, manganese or zinc in Lm iPGAM induces the formation of a mimic of the phosphoserine intermediate by the conformational change of Ser75 to interact with the phosphor group of the substrate/product. There are two metal sites located in the active site together with one molecule of the substrate/product in the crystal structures of Lm iPGAM. The dissociation constant of Co1 Lm iPGAM is much lower than that of Co2. The Kd of Co2 determined by activity assays is 9.1μM, whereas a crystallographic titration gives 0.28mM.
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