Study on the post-translational modification of RiPPs Xye catalyzed by CyFE PacB

HAN Yuanjun; MO Tianlu; DENG Zixin; ZHANG Qi; DING Wei

doi:10.12211/2096-8280.2021-080

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Study on the post-translational modification of RiPPs Xye catalyzed by CyFE PacB

Research Article | 更新时间：2026-03-18

- Study on the post-translational modification of RiPPs Xye catalyzed by CyFE PacB
- Synthetic Biology Journal Vol. 2, Issue 5, Pages: 826-836(2021)
- 作者机构：
  
  1.上海交通大学生命科学技术学院，微生物代谢国家重点实验室，上海 200240
  2.复旦大学化学系，上海 200243
- 作者简介：
- 基金信息：
- DOI：10.12211/2096-8280.2021-080
  CLC： Q936
- Received：01 August 2021，
  
  Revised：2021-08-12，
  
  Published：31 October 2021
- 稿件说明：
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韩沅均，莫天录，邓子新，张琪，丁伟. 环番合成酶PacB催化Xye类核糖体肽翻译后修饰研究［J］. 合成生物学， 2021， 2（5）： 826-836

HAN Yuanjun， MO Tianlu， DENG Zixin， ZHANG Qi， DING Wei. Study on the post-translational modification of RiPPs Xye catalyzed by CyFE PacB［J］. Synthetic Biology Journal， 2021， 2（5）： 826-836
韩沅均，莫天录，邓子新，张琪，丁伟. 环番合成酶PacB催化Xye类核糖体肽翻译后修饰研究［J］. 合成生物学， 2021， 2（5）： 826-836 DOI： 10.12211/2096-8280.2021-080.

HAN Yuanjun， MO Tianlu， DENG Zixin， ZHANG Qi， DING Wei. Study on the post-translational modification of RiPPs Xye catalyzed by CyFE PacB［J］. Synthetic Biology Journal， 2021， 2（5）： 826-836 DOI： 10.12211/2096-8280.2021-080.

摘要

近年来，一类含有分子内三残基环番结构的新型核糖体肽引起了广泛关注。这类化合物通常由一类

-腺苷甲硫氨酸自由基酶催化核心肽中芳香氨基酸的sp

碳和相邻第三位氨基酸残基侧链的sp

碳之间形成C—C或C—O键，这类酶被称为三残基环番合成酶（3-CyFEs）。目前发现Xye类核糖体肽天然产物生物合成基因簇中普遍含有此类三残基环番合成酶。本文报道从

Photorhabdus australis

DSM 17609中发现一个新的Xye类核糖体肽生物合成基因簇

pac

。异源表达和体外活性重建表明其生物合成基因簇编码的三残基环番合成酶PacB负责pacpeptide核心肽内的三个分子内环的生成。体内和体外实验结果初步表明PacB具有较高的底物容忍性，三个环番结构相互独立形成，但形成效率存在明显的差异，从而初步揭示了三个环番的形成顺序。本研究拓展了对Xye类核糖体肽化合物家族翻译后修饰以及三

残基环番合成酶催化功能的了解。

Abstract

Cyclic ribosomally synthesized and post-translationally modified peptides (RiPPs) with macrocycles derived from three amino acid residues have attracted intense interest in recent years. Biosynthesis of these peptides usually involves a radical (

)-adenosylmethionine (rSAM) enzyme that catalyzes the formation of a C—C or C—O bond between an aromatic carbon and a side chain carbon of another residue. Such enzymes are named three-residue cyclophane forming enzymes (3-CyFEs). The rSAM enzyme family is known as one of the largest enzyme super families

which consists of more than 22000 members. rSAM enzymes are widely found in all three life domains as one of the earliest biocatalysts on earth. A large amount of microbial genomic information shows that many RiPPs biosynthesis gene clusters contain rSAM enzymes. Recently

several 3-CyFEs from Xye gene clusters

which form three cyclizations on the precursor peptides

were reported. In this paper

we report a new Xye RiPP pacpeptide and its key biosynthetic enzyme PacB from

Photorhabdus australis

DSM 17609. We coexpressed the

pacA

and

pacB

Escherichia coli

and reconstituted the activity of heterologously expressed protein to investigate the bioactivity of 3-CyFEs PacB. The high-resolution mass spectrometry assay indicates that PacB indeed led to forming all three cyclophanes of pacpeptide. Additionally

the mutation of Asn/Argto Gly on the precursor peptide resulted in only two cyclophane was found

which demonstrated that three cyclization reactions were generated independently with different efficiency. PacB also catalyzed all three cyclophanes formation on the precursor peptide with Asn/Arg mutated to Ala. These results expand our understanding of 3-CyFEs chemistry in natural product biosynthesis and provide insights into the post-modification of cyclic RiPPs with three-residue cyclophanes.

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Biosynthesis and chemical synthesis of ribosomally synthesized and post-translationally modified peptides containing aminovinyl cysteine

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State Key Laboratory of Microbial Metabolism，School of Life Sciences and Biotechnology，Shanghai Jiao Tong University

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