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Title
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Crystal Structures of Quinolinate Synthase in Complex with a Substrate Analogue, the Condensation Intermediate, and Substrate-Derived Product.
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Authors
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A.Volbeda,
C.Darnault,
O.Renoux,
D.Reichmann,
P.Amara,
S.Ollagnier de Choudens,
J.C.Fontecilla-Camps.
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Ref.
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J Am Chem Soc, 2016,
138,
11802-11809.
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PubMed id
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Abstract
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The enzyme NadA catalyzes the synthesis of quinolinic acid (QA), the precursor
of the universal nicotinamide adenine dinucleotide (NAD) cofactor. Here, we
report the crystal structures of complexes between the Thermotoga maritima (Tm)
NadA K219R/Y107F variant and (i) the first intermediate (W) resulting from the
condensation of dihydroxyacetone phosphate (DHAP) with iminoaspartate and (ii)
the DHAP analogue and triose-phosphate isomerase inhibitor
phosphoglycolohydroxamate (PGH). In addition, using the TmNadA K219R/Y21F
variant, we have reacted substrates and obtained a crystalline complex between
this protein and the QA product. We also show that citrate can bind to both
TmNadA K219R and its Y21F variant. The W structure indicates that condensation
causes dephosphorylation. We propose that catalysis by the K219R/Y107F variant
is arrested at the W intermediate because the mutated protein is unable to
catalyze its aldo-keto isomerization and/or cyclization that ultimately lead to
QA formation. Intriguingly, PGH binds to NadA with its phosphate group at the
site where the carboxylate groups of W also bind. Our results shed significant
light on the mechanism of the reaction catalyzed by NadA.
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