Mechanisms of unusual enzymes in the biosynthesis of a copper-containing antibiotic

摘要 酶是强大的生物催化剂，可以被重新利用，以实现高效和可持续的合成 工业和医药化合物。确定新的酶催化反应和 了解它们的催化机理对于扩展生物催化工具箱是至关重要的。我们 最近发现了一种独特的含铜抗生素，氟菌素C，它是由 铜绿假单胞菌，多药耐药医院感染的主要原因。通过 在研究铜绿假单胞菌是如何产生氟菌素C的过程中，我们发现了一种不寻常的生物合成途径 涉及到新的酶化学。例如，一种新的血红素的两种铁依赖酶- 加氧酶样酶家族催化碳切除和氧化脱羧基 N-羟基化反应。腺苷琥珀酸裂解酶家族的两种裂解酶 分别生成和断裂碳-硫键的新活性。我们的化学药剂 经鉴定，扩大了这些酶家族的催化能力。我们假设 所述氟菌素C生物合成酶使用新的催化机制，并且可以被重新定位或 被设计来产生新的分子。为了检验这一假设，我们将使用以下组合 酶动力学、结构生物学、光谱学和合成化学工具，重点是 氟菌素C生物合成中的三种酶。对这些不同酶的研究将推动 对它们各自的酶家族如何催化不同的化学有基本的了解。 确定酶的机制和底物范围将使它们能够进一步 探索用于生物催化的用途。

Abstract Enzymes are powerful biocatalysts that may be repurposed for efficient and sustainable synthesis of industrial and pharmaceutical compounds. Identifying novel enzyme-catalyzed reactions and understanding their catalytic mechanisms is essential for expanding the biocatalytic toolbox. We have recently discovered a unique copper-containing antibiotic, fluopsin C, which is produced by Pseudomonas aeruginosa, a leading cause of multidrug-resistant nosocomial infections. By studying how P. aeruginosa makes fluopsin C, we identified an unusual biosynthetic pathway that involves novel enzyme chemistry. For instance, two iron-dependent enzymes of a new heme- oxygenase-like enzyme family catalyze a carbon excision and an oxidative decarboxylation and N-hydroxylation respectively. Two lyases of the adenylosuccinate lyase enzyme family exhibit novel activities of making and breaking carbon-sulfur bonds respectively. The chemistries we identified have expanded the catalytic capabilities of these enzyme families. We hypothesize that the fluopsin C biosynthetic enzymes use novel catalytic mechanisms and can be repurposed or engineered to generate new molecules. To test this hypothesis, we will use a combination of enzyme kinetics, structural biology, spectroscopy, and synthetic chemistry tools, with a focus on three enzymes in fluopsin C biosynthesis. Studies of these distinct enzymes will advance the fundamental understanding of how their respective enzyme family catalyzes diverse chemistry. Defining the mechanisms and substrate scope of the enzymes will allow them to be further explored for biocatalytic use.