Bifunctional NadC Homologue PyrZ Catalyzes Nicotinic Acid Formation in Pyridomycin Biosynthesis
Bifunctional NadC Homologue PyrZ Catalyzes Nicotinic Acid Formation in Pyridomycin Biosynthesis
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DOI：
10.1021/acschembio.2c00773
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发表时间：
2022-12-14
影响因子：
4
通讯作者：
Lin，Shuangjun
中科院分区：
文献类型：
--
作者：
Zhou，Zihua;Yang，Xu;Lin，Shuangjun
吡多霉素是一种有效的抗分枝杆菌天然产物，通过特异性抑制InhA，临床验证的抗结核药物发现目标。吡多霉素的吡啶基部分通过占据还原形式的烟酰胺腺嘌呤二核苷酸（NADH）辅因子结合位点在抑制InhA中发挥重要作用。在本文中，我们的生物化学特征PyrZ，这是一个多功能的NadC同系物和催化连续的形成，脱磷酸化，和核糖水解的烟酸monocluttide（NAMN），以产生烟酸（NA），生物合成的前体吡啶部分的pyridomycin。PyrZ与底物喹啉酸（QA）和最终产物NA的复合物的晶体结构揭示了K184与QA的C3-羧基之间形成的特异性盐桥。这种相互作用定位QA以接受磷酸核糖基以产生NAMN，将NAMN保留在活性位点内，并介导其转运至亲核体D296以进行去磷酸化。结合动力学和热力学分析以及定点突变，提出了PyrZ去磷酸化的催化机理。我们的研究发现了一个替代和简洁的NA生物合成途径，涉及一个独特的多功能酶。
Pyridomycin is a potent antimycobacterial natural product by specifically inhibiting InhA, a clinically validated antituberculosis drug discovery target. Pyridyl moieties of pyridomycin play an essential role in inhibiting InhA by occupying the reduced form of the nicotinamide adenine dinucleotide (NADH) cofactor binding site. Herein, we biochemically characterize PyrZ that is a multifunctional NadC homologue and catalyzes the successive formation, dephosphorylation, and ribose hydrolysis of nicotinic acid mononucleotide (NAMN) to generate nicotinic acid (NA), a biosynthetic precursor for the pyridyl moiety of pyridomycin. Crystal structures of PyrZ in complex with substrate quinolinic acid (QA) and the final product NA revealed a specific salt bridge formed between K184 and the C3-carboxyl group of QA. This interaction positions QA for accepting the phosphoribosyl group to generate NAMN, retains NAMN within the active site, and mediates its translocation to nucleophile D296 for dephosphorylation. Combining kinetic and thermodynamic analysis with site-directed mutagenesis, the catalytic mechanism of PyrZ dephosphorylation was proposed. Our study discovered an alternative and concise NA biosynthetic pathway involving a unique multifunctional enzyme.