Quinolinate synthase, also called NadA, is a [4Fe-4S]-containing enzyme that uses what is probably the oldest pathway to generate quinolinic acid (QA), the universal precursor of the biologically essential cofactor nicotinamide adenine dinucleotide (NAD). Its synthesis comprises the condensation of dihydroxyacetone phosphate (DHAP) and iminoaspartate (IA), which involves dephosphorylation, isomerization, cyclization, and two dehydration steps. The convergence of the three homologous domains of NadA defines a narrow active site that contains a catalytically essential [4Fe-4S] cluster. A tunnel, which can be opened or closed depending on the nature (or absence) of the bound ligand, connects this cofactor to the protein surface. One outstanding riddle has been the observation that the so far characterized active site is too small to bind IA and DHAP simultaneously. Here, we have used site-directed mutagenesis, X-ray crystallography, functional analyses, and molecular dynamics simulations to propose a condensation mechanism that involves the transient formation of a second active site cavity to which one of the substrates can migrate before this reaction takes place.

中文翻译：

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NadA合成喹啉酸过程中第二个活性位点腔的瞬态形成

喹啉酸合酶，也称为 NadA，是一种含有 [4Fe-4S] 的酶，它使用可能是最古老的途径来产生喹啉酸 (QA)，它是生物必需的辅因子烟酰胺腺嘌呤二核苷酸 (NAD) 的通用前体。其合成包括磷酸二羟丙酮 (DHAP) 和亚氨基天冬氨酸 (IA) 的缩合，包括脱磷酸、异构化、环化和两个脱水步骤。NadA 的三个同源结构域的收敛定义了一个狭窄的活性位点，其中包含一个催化必需的 [4Fe-4S] 簇。可以根据结合配体的性质（或不存在）打开或关闭的隧道将这个辅因子连接到蛋白质表面。一个突出的谜团是观察到迄今为止表征的活性位点太小而不能同时结合 IA 和 DHAP。在这里，我们使用定点诱变、X 射线晶体学、功能分析和分子动力学模拟来提出一种冷凝机制，该机制涉及瞬态形成第二个活性位点腔，其中一个底物可以在该反应发生之前迁移到该腔中。地方。