The Rut Pathway for Pyrimidine Degradation: Novel Chemistry and Toxicity Problems
The Rut Pathway for Pyrimidine Degradation: Novel Chemistry and Toxicity Problems
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DOI：
10.1128/jb.00201-10
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发表时间：
2010-08-01
影响因子：
3.2
通讯作者：
Wemmer, David E.
中科院分区：
文献类型：
--
作者：
Kim, Kwang-Seo;Pelton, Jeffrey G.;Wemmer, David E.
Rut途径由七种蛋白质组成，所有这些蛋白质都是大肠杆菌K-12在尿嘧啶作为唯一氮源上生长所必需的。RutA和RutB蛋白是核心：在缺乏它们的菌株中没有自发的抑制因子。RutA与黄素还原酶（RutF或替代物）一起催化一种新的反应。它直接裂解N-3和C-4之间的尿嘧啶环，产生脲基丙烯酸酯，如核磁共振（NMR）光谱和质谱所确定的。虽然脲基丙烯酸酯似乎是通过水解产生的，但反应的要求和从分子氧在C-4位引入O-18表明并非如此。质谱分析显示在反应混合物中存在少量具有脲基丙烯酸酯过酸质量的产物，并且我们推断这是RutA的直接产物。在体外RutB水解切割脲基丙烯酸酯，释放2摩尔的铵，丙二酸半醛，和二氧化碳。推测直接产物为氨基丙烯酸酯和氨基甲酸酯，两者均自发水解。结合生物信息学预测和已发表的晶体结构，遗传和生理学研究使我们能够预测RutC，-D和-E的功能。在体内，我们假设RutB水解脲基丙烯酸酯的过酸，产生氨基丙烯酸酯的过酸。我们推测，RutC减少氨基丙烯酸酯过酸氨基丙烯酸酯和RutD增加氨基丙烯酸酯的自发水解速率。吕特的功能似乎与YdfG相同，其将丙二酸半醛还原为3-羟基丙酸。RutG似乎是一种尿嘧啶转运蛋白。
The Rut pathway is composed of seven proteins, all of which are required by Escherichia coli K-12 to grow on uracil as the sole nitrogen source. The RutA and RutB proteins are central: no spontaneous suppressors arise in strains lacking them. RutA works in conjunction with a flavin reductase (RutF or a substitute) to catalyze a novel reaction. It directly cleaves the uracil ring between N-3 and C-4 to yield ureidoacrylate, as established by both nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry. Although ureidoacrylate appears to arise by hydrolysis, the requirements for the reaction and the incorporation of O-18 at C-4 from molecular oxygen indicate otherwise. Mass spectrometry revealed the presence of a small amount of product with the mass of ureidoacrylate peracid in reaction mixtures, and we infer that this is the direct product of RutA. In vitro RutB cleaves ureidoacrylate hydrolytically to release 2 mol of ammonium, malonic semialdehyde, and carbon dioxide. Presumably the direct products are aminoacrylate and carbamate, both of which hydrolyze spontaneously. Together with bioinformatic predictions and published crystal structures, genetic and physiological studies allow us to predict functions for RutC, -D, and -E. In vivo we postulate that RutB hydrolyzes the peracid of ureidoacrylate to yield the peracid of aminoacrylate. We speculate that RutC reduces aminoacrylate peracid to aminoacrylate and RutD increases the rate of spontaneous hydrolysis of aminoacrylate. The function of RutE appears to be the same as that of YdfG, which reduces malonic semialdehyde to 3-hydroxypropionic acid. RutG appears to be a uracil transporter.