Thioesterase-Catalyzed Aminoacylation and Thiolation of Polyketides in Fungi.
Thioesterase-Catalyzed Aminoacylation and Thiolation of Polyketides in Fungi.
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DOI：
10.1021/jacs.9b01083
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发表时间：
2019-05
影响因子：
15
通讯作者：
Mancheng Tang;Curt R. Fischer;Jason V. Chari;D. Tan;Sundari Suresh;A. Chu;Molly Miranda;Justin Smith;Zhuan Zhang;N. Garg;Robert P. St. Onge;Yi Tang
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文献类型：
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作者：
Mancheng Tang;Curt R. Fischer;Jason V. Chari;D. Tan;Sundari Suresh;A. Chu;Molly Miranda;Justin Smith;Zhuan Zhang;N. Garg;Robert P. St. Onge;Yi Tang
真菌高度还原聚酮合酶（HRPKS）使用一组结构域迭代地生物合成聚酮化合物。产品释放是 HRPKS 功能的关键步骤，以确保及时终止和酶周转。迄今为止，几乎所有 HRPKS 都采用单独的硫酯酶 (TE) 或酰基转移酶来释放产物。在本研究中，我们表征了两种具有融合 C 端 TE 结构域的真菌 HRPKS，这是真菌 HRPKS 的新结构域。我们表明，两种 HRPKS-TE 都以不依赖于 ATP 的方式合成氨酰化聚酮化合物。 KU42 TE 结构域选择半胱氨酸和同型半胱氨酸，并使用侧链硫醇基团作为亲核试剂催化转硫酯化反应。相比之下，KU43 TE 结构域选择亮氨酸甲酯并进行聚酮化合物的直接酰胺化，该反应通常由非核糖体肽合成酶 (NRPS) 结构域催化。这些 HRPKS-TE 酶的表征展示了 HRPKS 酶的功能多样性，并提供了潜在的 TE 结构域作为生物催化工具，使 HRPKS 结构多样化。
Fungal highly reducing polyketide synthases (HRPKSs) biosynthesize polyketides using a single set of domains iteratively. Product release is a critical step in HRPKS function to ensure timely termination and enzyme turnover. Nearly all of the HRPKSs characterized to date employ a separate thioesterase (TE) or acyltransferase enzyme for product release. In this study, we characterized two fungal HRPKSs that have fused C-terminal TE domains, a new domain architecture for fungal HRPKSs. We showed that both HRPKS-TEs synthesize aminoacylated polyketides in an ATP-independent fashion. The KU42 TE domain selects cysteine and homocysteine and catalyzes transthioesterification using the side-chain thiol group as the nucleophile. In contrast, the KU43 TE domain selects leucine methyl ester and performs a direct amidation of the polyketide, a reaction typically catalyzed by nonribosomal peptide synthetase (NRPS) domains. The characterization of these HRPKS-TE enzymes showcases the functional diversity of HRPKS enzymes and provides potential TE domains as biocatalytic tools to diversify HRPKS structures.