Biochemical and Structural Characterization of MycCI, a Versatile P450 Biocatalyst from the Mycinamicin Biosynthetic Pathway.
Biochemical and Structural Characterization of MycCI, a Versatile P450 Biocatalyst from the Mycinamicin Biosynthetic Pathway.
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MyCCI的生化和结构表征，MyCCI，一种来自霉菌素生物合成途径的多功能P450生物催化剂。
DOI：
10.1021/acschembio.6b00479
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发表时间：
2016-09-16
影响因子：
4
通讯作者：
Sherman, David H.
中科院分区：
文献类型：
--
作者：
DeMars, Matthew D., II;Sheng, Fang;Park, Sung Ryeol;Lowell, Andrew N.;Podust, Larissa M.;Sherman, David H.
细胞色素P450单加氧酶(P450)是自然界中最普遍和最多功能的酶，用于进行氧化代谢转化。它们选择性地使惰性C-H键功能化的能力无与伦比，这导致它们在学术和工业环境中越来越多地用于生产精细和商品化学品。许多最有趣和潜在的生物催化有用的P450来自微生物，在那里它们催化天然产品生物合成途径中的关键剪裁反应。虽然这些酶大多作用于结构复杂的途径中间体，具有很高的选择性，但它们的底物范围往往很窄，从而限制了它们的广泛应用。在本研究中，我们研究了从霉素生物合成途径中的P450 MycCI对各种大环化合物的反应性，发现该酶对几个16元环大内酯表现出明显的活性，而不依赖于它们的糖基化状态。这些结果得到了平衡底物结合实验、稳态动力学研究和MycCI与其天然底物Mycinamicin VIII结合的X射线结晶学分析的证实。我们还对泰乐菌素途径的同源P450 TylHI进行了表征，表明其底物范围比MycCI受到严重限制。因此，后者能够羟化大环苷元和大环内酯类化合物，使其有别于相关的生物合成P450，并突出了其开发具有广泛底物范围和高区域选择性的新型P450生物催化剂的潜力。
Cytochrome P450 monooxygenases (P450s) are some of nature’s most ubiquitous and versatile enzymes for performing oxidative metabolic transformations. Their unmatched ability to selectively functionalize inert C-H bonds has led to their increasing employment in academic and industrial settings for the production of fine and commodity chemicals. Many of the most interesting and potentially biocatalytically useful P450s come from microorganisms, where they catalyze key tailoring reactions in natural product biosynthetic pathways. While most of these enzymes act on structurally complex pathway intermediates with high selectivity, they often exhibit narrow substrate scope, thus limiting their broader application. In the present study, we investigated the reactivity of the P450 MycCI from the mycinamicin biosynthetic pathway toward a variety of macrocyclic compounds and discovered that the enzyme exhibits appreciable activity on several 16-membered ring macrolactones independent of their glycosylation state. These results were corroborated by performing equilibrium substrate binding experiments, steady-state kinetics studies, and x-ray crystallographic analysis of MycCI bound to its native substrate mycinamicin VIII. We also characterized TylHI, a homologous P450 from the tylosin pathway, and showed that its substrate scope is severely restricted compared to MycCI. Thus, the ability of the latter to hydroxylate both macrocyclic aglycones and macrolides sets it apart from related biosynthetic P450s and highlights its potential for developing novel P450 biocatalysts with broad substrate scope and high regioselectivity.
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影响因子：
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