Journal
MOLECULAR AND BIOCHEMICAL PARASITOLOGY
Volume 157, Issue 2, Pages 179-186Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.molbiopara.2007.10.013
Keywords
giardia; species-specific inhibition; thiol reagents; triosephosphate isomerase
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Giardia lamblia depends on glycolysis to obtain ATP, highlighting the suitability of glycolytic enzymes as targets for drug design. We studied triosephosphate isomerase from G. lamblia (GITIM) as a potential species-specific drug target. Cysteine-reactive agents were used as probes, in order to test those regions near to cysteine residues as targets to perturb enzyme structure and activity. Methyl methanethiosulfonate (MMTS) derivatized three of the five Cys per subunit of dimeric GITIM and induced 50% of inactivation. The 2-carboxyethyl methanethiosulfonate (MTSCE) modified four Cys and induced 97% of inactivation. Inactivation by MMTS or MTSCE did not affect secondary structure, nor induce dimer dissociation; however, Cys modification decreased thermal stability of enzyme. Inactivation and dissociation of the dimer to stable monomers were reached when four Cys were derivatized by 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNB). The effects of DTNB were completely abolished when GITIM was first treated with MMTS. The effect of thiol reagents on human TIM was also assayed; it is 180-fold less sensitive than GITIM. Collectively, the data illustrate GITIM as a good target for drug design. (c) 2007 Elsevier B.V. All rights reserved.
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