Structural and functional insight into an unexpectedly selective N-methyltransferase involved in plantazolicin biosynthesis
Structural and functional insight into an unexpectedly selective N-methyltransferase involved in plantazolicin biosynthesis
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DOI：
10.1073/pnas.1306101110
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发表时间：
2013-08-06
影响因子：
11.1
通讯作者：
Mitchell, Douglas A.
中科院分区：
文献类型：
--
作者：
Lee, Jaeheon;Hao, Yue;Mitchell, Douglas A.
Plantazolicin (PZN) 是一种多杂环 N-α,N-α-二甲基精氨酸抗生素，对炭疽杆菌（炭疽杆菌）菌株具有显着的特异性杀菌活性。先前的研究表明，从 PZN 生物合成基因簇中遗传删除 S-腺苷-L-甲硫氨酸依赖性甲基转移酶会导致去甲基 PZN 的形成，而去甲基 PZN 缺乏抗生素活性。在这里，我们描述了 PZN 甲基转移酶的体外重建、突变分析和 X 射线晶体结构。与所有其他已知的在体外作用于不同底物的小分子甲基转移酶不同，PZN 甲基转移酶的底物范围异常有限，并且仅对去甲基 PZN 及其相近衍生物起作用。两种相关的 PZN 甲基转移酶的晶体结构，分辨率为 1.75 埃（解淀粉芽孢杆菌）和 2.0 埃（短小芽孢杆菌），揭示了一个又深又窄的空腔，推测其作为去甲基 PZN 的结合位点。该腔的狭窄性为理解极端底物选择性的分子基础提供了一个框架。对解淀粉芽孢杆菌甲基转移酶的一组点突变进行分析，可以鉴定具有结构和催化重要性的残基。这些发现进一步加深了我们对涉及噻唑/恶唑修饰的小菌素生物合成的一组直系同源酶的理解，这是一个快速发展的天然产物研究领域。
Plantazolicin (PZN), a polyheterocyclic, N-alpha, N-alpha-dimethylarginine-containing antibiotic, harbors remarkably specific bactericidal activity toward strains of Bacillus anthracis, the causative agent of anthrax. Previous studies demonstrated that genetic deletion of the S-adenosyl-L-methionine-dependent methyltransferase from the PZN biosynthetic gene cluster results in the formation of desmethylPZN, which is devoid of antibiotic activity. Here we describe the in vitro reconstitution, mutational analysis, and X-ray crystallographic structure of the PZN methyltransferase. Unlike all other known small molecule methyltransferases, which act upon diverse substrates in vitro, the PZN methyltransferase is uncharacteristically limited in substrate scope and functions only on desmethylPZN and close derivatives. The crystal structures of two related PZN methyltransferases, solved to 1.75 angstrom (Bacillus amyloliquefaciens) and 2.0 angstrom (Bacillus pumilus), reveal a deep, narrow cavity, putatively functioning as the binding site for desmethylPZN. The narrowness of this cavity provides a framework for understanding the molecular basis of the extreme substrate selectivity. Analysis of a panel of point mutations to the methyltransferase from B. amyloliquefaciens allowed the identification of residues of structural and catalytic importance. These findings further our understanding of one set of orthologous enzymes involved in thiazole/oxazole-modified microcin biosynthesis, a rapidly growing sector of natural products research.