Molecular insights into the enzymatic diversity of flavin-trafficking protein (Ftp; formerly ApbE) in flavoprotein biogenesis in the bacterial periplasm.
Molecular insights into the enzymatic diversity of flavin-trafficking protein (Ftp; formerly ApbE) in flavoprotein biogenesis in the bacterial periplasm.
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DOI：
10.1002/mbo3.306
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发表时间：
2016-02
期刊：
影响因子：
3.4
通讯作者：
Norgard MV
中科院分区：
文献类型：
--
作者：
Deka RK;Brautigam CA;Liu WZ;Tomchick DR;Norgard MV
我们最近报道了梅毒螺旋体梅毒螺旋体（Ftp_Tp）中的黄素运输蛋白（Ftp），它是第一个细菌金属依赖性 FAD 焦磷酸酶，可将周质中的 FAD 水解为 AMP 和 FMN。其他细菌中 Ftp_Tp 的直系同源物（以前称为 ApbE）似乎缺乏这种水解活性；相反，它们通过金属依赖性 FMN 转移酶活性使氧化还原亚基 NqrC 黄素酰化。然而，对于含 Nqr 或 Rnf 氧化还原细菌中金属依赖性 Ftp 催化的性质或机制尚不清楚。在当前的研究中，我们在大肠杆菌 Ftp (Ftp_Ec) 晶体结构中发现了一个双金属中心，并通过诱变表明，单个氨基酸取代可将其从 FAD 结合蛋白转化为 Mg2+ 依赖性 FAD 焦磷酸酶（Ftp_Tp 类）。此外，在蛋白质底物存在的情况下，两种类型的 Ftps 都能够通过 FMN 的金属依赖性共价连接使周质氧化还原蛋白（例如 RnfG_Ec）黄素化。 Shewanella oneidensis NqrC 的 Ftp 介导的黄素酰化蛋白的高分辨率结构鉴定出翻译后修饰的黄素蛋白中磷酸酯-苏氨酰-FMN 键形成中必需的赖氨酸。总之，这些发现拓宽了我们对细菌周质生理能力的理解，并且还阐明了黄素蛋白产生的可能机制。
We recently reported a flavin‐trafficking protein (Ftp) in the syphilis spirochete Treponema pallidum (Ftp_Tp) as the first bacterial metal‐dependent FAD pyrophosphatase that hydrolyzes FAD into AMP and FMN in the periplasm. Orthologs of Ftp_Tp in other bacteria (formerly ApbE) appear to lack this hydrolytic activity; rather, they flavinylate the redox subunit, NqrC, via their metal‐dependent FMN transferase activity. However, nothing has been known about the nature or mechanism of metal‐dependent Ftp catalysis in either Nqr‐ or Rnf‐redox‐containing bacteria. In the current study, we identified a bimetal center in the crystal structure of Escherichia coli Ftp (Ftp_Ec) and show via mutagenesis that a single amino acid substitution converts it from an FAD‐binding protein to a Mg2+‐dependent FAD pyrophosphatase (Ftp_Tp‐like). Furthermore, in the presence of protein substrates, both types of Ftps are capable of flavinylating periplasmic redox‐carrying proteins (e.g., RnfG_Ec) via the metal‐dependent covalent attachment of FMN. A high‐resolution structure of the Ftp‐mediated flavinylated protein of Shewanella oneidensis NqrC identified an essential lysine in phosphoester‐threonyl‐FMN bond formation in the posttranslationally modified flavoproteins. Together, these discoveries broaden our understanding of the physiological capabilities of the bacterial periplasm, and they also clarify a possible mechanism by which flavoproteins are generated.