Characterization of a Hybrid Nonribosomal Peptide-Carbohydrate Biosynthetic Pathway in Photorhabdus luminescens.
Characterization of a Hybrid Nonribosomal Peptide-Carbohydrate Biosynthetic Pathway in Photorhabdus luminescens.
复制标题
发光杆菌中混合非核糖体肽-碳水化合物生物合成途径的表征。
DOI：
10.1021/acs.biochem.8b01120
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发表时间：
2019
期刊：
影响因子：
2.9
通讯作者：
Crawford,JasonM
中科院分区：
文献类型：
--
作者：
Perez,CoreyE;Crawford,JasonM
基因组测序和分析的进步提供了大量“孤儿”细菌生物合成途径，其中许多含有假设的蛋白质。鉴于这些假设的蛋白质具有进行新化学反应的潜力，孤儿途径可以作为发现新酶的丰富库，这些新酶负责产生具有令人着迷的化学和生物功能的代谢物。我们之前在昆虫病原体发光杆菌中发现了一种罕见的混合非核糖体肽合成酶（NRPS）-碳水化合物基因组岛。该途径的异源表达导致了含有 1,6-脱水-β-d-N-乙酰基-葡萄糖胺部分的寡糖的表征，但这些新的代谢物缺乏 NRPS 机制的修饰。在这里，通过应用自上而下的蛋白质质谱、途径靶向分子网络、稳定同位素标记和体外蛋白质生物化学，我们完成了该生物合成途径的表征，并鉴定了该途径的混合产物，一种新的“糖氨基酸”代谢物，称为光糖。有趣的是，一种假设的蛋白质充当桥梁，将源自 NRPS 机制的甘氨酰单元缩合到游离的 1,6-脱水-β-d-N-乙酰基-葡萄糖胺核心上。我们进一步证明该基因簇赋予抗菌肽挑战生长优势。
Advances in genome sequencing and analysis have afforded a trove of “orphan” bacterial biosynthetic pathways, many of which contain hypothetical proteins. Given the potential for these hypothetical proteins to carry out novel chemistry, orphan pathways serve as a rich reservoir for the discovery of new enzymes responsible for the production of metabolites with both fascinating chemistries and biological functions. We previously identified a rare hybrid nonribosomal peptide synthetase (NRPS)–carbohydrate genomic island in the entomopathogenPhotorhabdus luminescens. Heterologous expression of the pathway led to the characterization of oligosaccharides harboring a 1,6-anhydro-β-d-N-acetyl-glucosamine moiety, but these new metabolites lacked modification by the NRPS machinery. Here, through the application of top-down protein mass spectrometry, pathway-targeted molecular networking, stable isotope labeling, andin vitroprotein biochemistry, we complete the characterization of this biosynthetic pathway and identify the hybrid product of the pathway, a new “glycoamino acid” metabolite termed photolose. Intriguingly, a hypothetical protein served as a bridge to condense a glycyl unit derived from the NRPS machinery onto the free 1,6-anhydro-β-d-N-acetyl-glucosamine core. We further demonstrate that the gene cluster confers a growth advantage to antimicrobial peptide challenge.