Rational design, synthesis and evaluation of first generation inhibitors of the Giardia lamblia fructose-1,6-biphosphate aldolase |
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Authors: | Li Zhimin Liu Zhengang Cho Dae Won Zou Jiwen Gong Maozhen Breece Robert M Galkin Andrey Li Ling Zhao Hong Maestas Gabriel D Tierney David L Herzberg Osnat Dunaway-Mariano Debra Mariano Patrick S |
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Institution: | a Department of Chemistry and Chemical Biology, University of New Mexico, Albuquerque, NM 87131, United Statesb Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056, United Statesc Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute, Rockville, MD 20850, United States |
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Abstract: | Inhibitors of the Giardia lamblia fructose 1,6-bisphosphate aldolase (GlFBPA), which transforms fructose 1,6-bisphosphate (FBP) to dihydroxyacetone phosphate and glyceraldehyde 3-phosphate, were designed based on 3-hydroxy-2-pyridone and 1,2-dihydroxypyridine scaffolds that position two negatively charged tetrahedral groups for interaction with substrate phosphate binding residues, a hydrogen bond donor to the catalytic Asp83, and a Zn2+ binding group. The inhibition activities for the GlFBPA catalyzed reaction of FBP of the prepared alkyl phosphonate/phosphate substituted 3-hydroxy-2-pyridinones and a dihydroxypyridine were determined. The 3-hydroxy-2-pyridone inhibitor 8 was found to bind to GlFBPA with an affinity (Ki = 14 μM) that is comparable to that of FBP (Km = 2 μM) or its inert analog TBP (Ki = 1 μM). The X-ray structure of the GlFBPA-inhibitor 8 complex (2.3 Å) shows that 8 binds to the active site in the manner predicted by in silico docking with the exception of coordination with Zn2+. The observed distances and orientation of the pyridone ring O=C-C-OH relative to Zn2+ are not consistent with a strong interaction. To determine if Zn2+coordination occurs in the GlFBPA-inhibitor 8 complex in solution, EXAFS spectra were measured. A four coordinate geometry comprised of the three enzyme histidine ligands and an oxygen atom from the pyridone ring O=C-C-OH was indicated. Analysis of the Zn2+ coordination geometries in recently reported structures of class II FBPAs suggests that strong Zn2+ coordination is reserved for the enediolate-like transition state, accounting for minimal contribution of Zn2+ coordination to binding of 8 to GlFBPA. |
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Keywords: | Fructose-1 6-biphosphate aldolase Giardia lamblia Zn2+ coordination Hydroxypyridinone EXAFS Inhibitor design |
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