Mapping and Exploiting the Promiscuity of OxyB toward the Biocatalytic Production of Vancomycin Aglycone Variants.
Mapping and Exploiting the Promiscuity of OxyB toward the Biocatalytic Production of Vancomycin Aglycone Variants.
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DOI：
10.1021/acscatal.0c01719
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发表时间：
2020-08-21
期刊：
影响因子：
12.9
通讯作者：
Seyedsayamdost MR
中科院分区：
文献类型：
--
作者：
Forneris CC;Nguy AKL;Seyedsayamdost MR
万古霉素是临床对抗传染病最重要的抗生素之一。其生物活性依赖于三个芳香族交联，这些交联形成杯形拓扑并允许与新生肽聚糖链紧密结合。细胞色素 P450 酶 OxyB、OxyA 和 OxyC 已被证明可以引入这些合成上具有挑战性的芳香键。在化学酶方案中利用 P450 酶生成万古霉素衍生物的能力很有吸引力，但需要对其反应性和机制有透彻的了解。在此，我们系统地探索了 OxyB 生物催化的范围，并报告了当酶与修饰的万古霉素前体肽一起存在时，具有不同大环尺寸和组成的多种二芳基醚和联芳基交联的安装。使用一维/二维核磁共振光谱、高分辨率质谱（HR-MS）、串联HR-MS和同位素标记以及紫外-可见光吸收和荧光发射光谱确定了所得产物的结构。对这些替代 OxyB 产品的生物活性的探索令人惊讶地揭示了抗真菌特性。利用 OxyB 的混杂性，我们通过化学酶法生成了包含扩展大环的万古霉素糖苷配基变体。讨论了 OxyB 的机制意义以及创建万古霉素类似物库的未来方向。
Vancomycin is one of the most important clinical antibiotics in the fight against infectious disease. Its biological activity relies on three aromatic cross-links, which create a cup-shaped topology and allow tight binding to nascent peptidoglycan chains. The cytochrome P450 enzymes OxyB, OxyA, and OxyC have been shown to introduce these synthetically challenging aromatic linkages. The ability to utilize the P450 enzymes in a chemo-enzymatic scheme to generate vancomycin derivatives is appealing but requires a thorough understanding of their reactivities and mechanisms. Herein, we systematically explore the scope of OxyB biocatalysis and report installation of diverse diaryl ether and biaryl cross-links with varying macrocycle sizes and compositions, when the enzyme is presented with modified vancomycin precursor peptides. The structures of the resulting products were determined using one-dimensional/two-dimensional nuclear magnetic resonance spectroscopy, high-resolution mass spectrometry (HR-MS), tandem HR-MS, and isotopic labeling, as well as ultraviolet–visible light absorption and fluorescence emission spectroscopies. An exploration of the biological activities of these alternative OxyB products surprisingly revealed antifungal properties. Taking advantage of the promiscuity of OxyB, we chemo-enzymatically generated a vancomycin aglycone variant containing an expanded macrocycle. Mechanistic implications for OxyB and future directions for creating vancomycin analogue libraries are discussed.
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