Du Yanan, Qiu Yanping, Meng Xiang, Feng Junyin, Tao Jiang, Liu Wen
State Key Laboratory of Bioorganic and Natural Products Chemistry, Center for Excellence in Molecular Synthesis, Shanghai Institute of Organic Chemistry, University of Chinese Academy of Sciences, 345 Lingling Road, Shanghai 200032, China.
Huzhou Center of Bio-Synthetic Innovation, 1366 Hongfeng Road, Huzhou 313000, China.
J Am Chem Soc. 2020 May 6;142(18):8454-8463. doi: 10.1021/jacs.0c02329. Epub 2020 Apr 24.
Sulfomycins are sulfur-rich, ribosomally synthesized, and post-translationally modified peptides (RiPPs) that are characterized by a 35-membered macrocyclic ring system with a pyridine domain central to five azoles and additional dehydroamino acids. The pathway through which these large thiopeptide antibiotics are formed in remains elusive. Starting with the cloning of the biosynthetic gene cluster of sulfomycins, we here dissect a two-stage process in which an unusual dehydrogenase heterotrimer functions with two distinct YcaO proteins to install five azole heterocycles into the core peptide sequence of the precursor peptide. The first stage involves the activity of a typical cyclodehydratase complex composed of a didomain E1-YcaO protein and an F-protein partner to heterocyclize distant residues l-Cys2 and l-Thr9 and then the activity of the heterotrimeric dehydrogenase complex that converts the resulting azolines to azoles. In the second stage, this dehydrogenase complex associates with a discrete YcaO protein to form an atypical, four-component azole synthase complex, which is capable of sequentially converting residues l-Cys7, l-Thr5, and l-Ser12 to azoles in a distinct manner. During this process, an E1-like partner protein plays a critical role and functions through the two stages to mediate a variety of specific protein-protein interactions. This partner protein participates in the formation of the active dehydrogenase heterotrimer and the engagement of discrete YcaO activity to form the azole synthase heterotetramer. The findings in this study advance the understanding in the biosynthesis of different azole-containing RiPPs and set the stage for the discovery, engineering, and creation of new thiopeptides using genome mining and synthetic biology approaches.
硫霉素是富含硫的、核糖体合成并经翻译后修饰的肽（RiPPs），其特征在于具有一个35元大环系统，该系统带有一个吡啶结构域，该结构域位于五个唑的中心位置以及额外的脱氢氨基酸。这些大型硫肽抗生素在体内形成的途径仍然不清楚。从硫霉素生物合成基因簇的克隆开始，我们在此剖析了一个两阶段过程，其中一个不寻常的脱氢酶异源三聚体与两种不同的YcaO蛋白协同作用，将五个唑杂环安装到前体肽的核心肽序列中。第一阶段涉及由双结构域E1 - YcaO蛋白和F蛋白伴侣组成的典型环化脱水酶复合物的活性，以使远距离的残基l - Cys2和l - Thr9杂环化，然后是异源三聚体脱氢酶复合物的活性，该复合物将生成的唑啉转化为唑。在第二阶段，该脱氢酶复合物与一个离散的YcaO蛋白结合，形成一个非典型的四组分唑合酶复合物，该复合物能够以独特的方式依次将残基l - Cys7、l - Thr5和l - Ser12转化为唑。在此过程中，一个类似E1的伴侣蛋白起着关键作用，并在两个阶段发挥功能，以介导各种特定的蛋白质 -蛋白质相互作用。该伴侣蛋白参与活性脱氢酶异源三聚体的形成以及离散YcaO活性的参与，以形成唑合酶异源四聚体。本研究中的发现推进了对不同含唑RiPPs生物合成的理解，并为使用基因组挖掘和合成生物学方法发现、工程改造和创造新的硫肽奠定了基础。
