Peptide cyclization catalysed by the thioesterase domain of tyrocidine synthetase
Peptide cyclization catalysed by the thioesterase domain of tyrocidine synthetase
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DOI：
10.1038/35025116
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发表时间：
2000-09-14
期刊：
影响因子：
64.8
通讯作者：
Walsh, CT
中科院分区：
文献类型：
--
作者：
Trauger, JW;Kohli, RM;Walsh, CT
在许多大环天然产物通过多结构域大合成酶的生物合成中，羧基末端硫酯酶（TE）结构域参与环化和产物释放（1，2）;然而，尚未确定TE结构域是否可以独立于上游结构域催化大环化（以及对称环肽的延伸）。不能将TE环化步骤与早期的链组装步骤分离已经排除了TE底物特异性的测定，这对于新化合物的工程化生物合成是重要的（1）。在这里，我们报告了从tyrocidine合成酶切下的TE结构域有效地催化十肽硫酯的环化形成抗生素tyrocidine A，并且可以催化五肽硫酯二聚化，然后环化形成抗生素短杆菌肽S。通过系统地改变十肽硫酯底物和比较环化速率，我们还表明，只有两个残基（一个附近的十肽的每一端）是环化的关键。这种特异性特征表明，酪氨酸合成酶TE，并且通过类推，许多其他TE结构域，将能够环化和释放由工程化酶组装线产生的广泛的新底物和产物。
In the biosynthesis of many macrocyclic natural products by multidomain megasynthases, a carboxy-terminal thioesterase (TE) domain is involved in cyclization and product release(1,2); however, it has not been determined whether TE domains can catalyse macrocyclization (and elongation in the case of symmetric cyclic peptides) independently of upstream domains. The inability to decouple the TE cyclization step from earlier chain assembly steps has precluded determination of TE substrate specificity, which is important for the engineered biosynthesis of new compounds(1). Here we report that the excised TE domain from tyrocidine synthetase efficiently catalyses cyclization of a decapeptide-thioester to form the antibiotic tyrocidine A, and can catalyse pentapeptide-thioester dimerization followed by cyclization to form the antibiotic gramicidin S. By systematically varying the decapeptide-thioester substrate and comparing cyclization rates, we also show that only two residues (one near each end of the decapeptide) are critical for cyclization. This specificity profile indicates that the tyrocidine synthetase TE, and by analogy many other TE domains, will be able to cyclize and release a broad range of new substrates and products produced by engineered enzymatic assembly lines.