Cyclization of fungal nonribosomal peptides by a terminal condensation-like domain.
Cyclization of fungal nonribosomal peptides by a terminal condensation-like domain.
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DOI：
10.1038/nchembio.1047
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发表时间：
2012-10
影响因子：
14.8
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中科院分区：
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由非核糖体肽合成酶（NRPSs）产生的线性肽基前体的环化是生物合成具有生物活性的环肽的重要步骤。细菌NRPSs使用硫酯酶（TE）结构域进行环化，而真菌NRPSs显然已经进化到使用不同的酶途径。在经过验证的产生大环肽的真菌NRPSs中，每个大合成酶以可能进行大环化反应的冷凝样（CT）结构域终止。为了探讨这种CT结构域的作用，我们在酿酒酵母和体外重建了埃塞俄比亚青霉三模NPRS TqaA的活性。与重组的双模NRPS AnaPS一起，我们剖析了TqaA在将线性邻氨基苯甲酸-d -色氨酸- l -丙氨酰三肽转化为烟喹唑啉f的环化步骤。大量的生化和突变研究证实了CT结构域在催化巯基化结构域依赖的环化过程中的重要作用。我们的工作提供了真菌NRPSs可能采用普遍大环化策略的证据。
Cyclization of linear peptidyl precursors produced by nonribosomal peptide synthetases (NRPSs) is an important step in the biosynthesis of bioactive cyclic peptides. Whereas bacterial NRPSs use thioesterase (TE) domains to perform the cyclization, fungal NRPSs have apparently evolved to use a different enzymatic route. In verified fungal NRPSs that produce macrocyclic peptides, each megasynthetase terminates with a condensation-like (CT) domain that may perform the macrocyclization reaction. To probe the role of such a CT domain, we reconstituted the activities of the Penicillium aethiopicum trimodular NPRS TqaA in Saccharomyces cerevisiae and in vitro. Together with a reconstituted bimodular NRPS AnaPS, we dissected the cyclization steps of TqaA in transforming the linear anthranilate-D-tryptophan-L-alanyl tripeptide into fumiquinazoline F. Extensive biochemical and mutational studies confirmed the essential role of the CT domain in catalyzing cyclization in a thiolation domain-dependent fashion. Our work provided evidence of a likely universal macrocyclization strategy employed by fungal NRPSs.