Characterization of the macrocyclase involved in the biosynthesis of RiPP cyclic peptides in plants
Characterization of the macrocyclase involved in the biosynthesis of RiPP cyclic peptides in plants
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DOI：
10.1073/pnas.1620499114
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发表时间：
2017-06-20
影响因子：
11.1
通讯作者：
Nair, Satish K.
中科院分区：
文献类型：
--
作者：
Chekan, Jonathan R.;Estrada, Paola;Nair, Satish K.
长期以来，人们一直在寻找能够催化线性多肽底物大环化的酶，以通过组合基因组装来生产结构多样化的支架文库，并提供快速的体内筛选方法。环状多肽是植物核糖体合成和翻译后修饰的多肽，具有不同的大小和拓扑结构，其中一些被证明具有生物活性。环肽在大小和序列上的多样性表明，相应的大环酶可能是生产环肽的理想催化剂。在这里，我们介绍了参与环状环化的植物酶PCY1的生化特性和晶体结构。这些研究表明，PCY1 S9A蛋白酶折叠已被适应于酰基酶中间体的转酰胺化，而不是水解化，以产生环状产物。值得注意的是，PCY1使用了一种不寻常的策略，即从底物上切割的C末端跟随肽稳定酶的有效构象，以促进N末端片段的大环化。PCY1广泛的底物耐受性可以作为一种生物技术工具来产生结构多样化的大环阵列，包括那些含有非蛋白生成元件的大环。
Enzymes that can catalyze the macrocyclization of linear peptide substrates have long been sought for the production of libraries of structurally diverse scaffolds via combinatorial gene assembly as well as to afford rapid in vivo screening methods. Orbitides are plant ribosomally synthesized and posttranslationally modified peptides (RiPPs) of various sizes and topologies, several of which are shown to be biologically active. The diversity in size and sequence of orbitides suggests that the corresponding macrocyclases may be ideal catalysts for production of cyclic peptides. Here we present the biochemical characterization and crystal structures of the plant enzyme PCY1 involved in orbitide macrocyclization. These studies demonstrate how the PCY1 S9A protease fold has been adapted for transamidation, rather than hydrolysis, of acyl-enzyme intermediates to yield cyclic products. Notably, PCY1 uses an unusual strategy in which the cleaved C-terminal follower peptide from the substrate stabilizes the enzyme in a productive conformation to facilitate macrocyclization of the N-terminal fragment. The broad substrate tolerance of PCY1 can be exploited as a biotechnological tool to generate structurally diverse arrays of macrocycles, including those with nonproteinogenic elements.