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Carine Rousset

Structural and functional studies of Quinolinate synthase: An iron-sulfur protein as a key target of antibacterial agents



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Published on 4 May 2009


Thesis presented May 04, 2009

Abstract:
Quinolinate synthase (NadA) catalyzes a unique condensation reaction between iminoaspartate and dihydroxacetone phosphate leading to quinolinic acid, a key intermediate in the biosynthesis of nicotinamide adenine dinucleotide (NAD). This study demonstrates that all quinolinate synthases harbor a [4Fe-4S] cluster, absolutely required for activity, which is coordinated by only three cysteine residues, namely the conserved Cys113, Cys200 and Cys297 in NadA from E. coli. Spectroscopic and biochemical properties of [4Fe-4S] clusters lead to the proposal that the iron-sulfur cluster plays an aconitase/dehydratase-type role in this enzymatic catalysis. Two other cysteine residues (Cys291 and Cys294 in E. coli) were demonstrated to be involved in the formation of a disulfide bridge critical for the quinolinate synthase activity probably by playing a regulatory role. In this work we also propose an additional step in the mechanism including the isomerization​ of glyceraldehyde 3-phosphate (G-3P) to DHAP. Finally, some putative inhibitory molecules were assayed on NadA that we consider as a potential target of antibacterial agents. Among the tested molecules, phosphoglycolohydroxamic acid (PGH) proves to be active on the quinolinate synthases of E. coli and M. tuberculosis, acting as a competitive inhibitor of DHAP.

Keywords:
Quinolinate synthase, Nicotinamide adenine dinucleotide (NAD), Dihydroxyacetone Phosphate, Iminoaspartate, quinolinic acid, [4Fe-4S], antibacterial agents, phosphoglycolohydroxamique acid (PGH)

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