Elucidating and exploiting halogenase recruitment beta hairpin docking domains in nonribosomal peptide biosynthesis

阐明和利用非核糖体肽生物合成中卤化酶募集β发夹对接结构域

• 批准号: 1782570
• 金额: --
• 依托单位: University of Warwick
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2016
• 资助国家: 英国
• 起止时间: 2016 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-1782570
• 关键词: Elucidating; exploiting; halogenase; recruitment; beta

Nonribosomal peptides and polyketides are classes of natural products, comprising a large variety of biologically-relevant compounds. They are synthesised by nonribosomal peptide synthases (NRPSs) and polyketide synthases (PKSs) respectively, which are modular megasynthases that act in an assembly line-like manner. The interaction between subunits of PKS and NRPS proteins is critical to the fidelity of their biosyntheses, and are known in many cases to be facilitated by C-terminal and N-terminal docking domains. One type of interaction between subunits in hybrid NRPS-PKS systems involves the docking of -hairpin docking domains (HDDs) attached to catalytic domains at the N-terminus of one subunit with a short linear motifs (SLiMs) attached to carrier proteins at the C-terminus of another subunit. Recent computational work in the Challis group has suggested that this system of docking domains may be far more prevalent in the biosynthetic machinery of polyketide and nonribosomal peptide metabolites than first appreciated. This project will focus upon the characterisation of HDD/SLiM interactions predicted to occur in the biosynthetic pathway of nonribosomal peptides from cyanobacteria. The aims of the project are to experimentally characterise the HDD/SLiM interactions by overproducing the relevant proteins in E. coli and using synthetic substrate analogues to examine their enzymatic activity. We will also attempt to obtain high resolution structural information for the proteins using X-ray crystallography and NMR spectroscopy. Once characterised, this system will be exploited to understand the potential of the HDD/SLiM systems as a tool for engineering of biosynthetic pathways, including the production of hybrid assembly lines using components from the biosyntheses of different metabolites.

非核糖体肽和聚酮化合物是一类天然产物，包含多种生物学相关化合物。它们分别由非核糖体肽合成酶（NRPS）和聚酮合成酶（PKS）合成，所述非核糖体肽合成酶和聚酮合成酶是以装配线样方式起作用的模块化大合成酶。PKS和NRPS蛋白亚基之间的相互作用对于其生物合成的保真度至关重要，并且已知在许多情况下通过C-末端和N-末端对接结构域来促进。杂合NRPS-PKS系统中亚基之间的一种类型的相互作用涉及在一个亚基的N-末端处连接到催化结构域的发夹对接结构域（HDD）与在另一个亚基的C-末端处连接到载体蛋白的短线性基序（SLiM）的对接。查利斯小组最近的计算工作表明，这种对接结构域系统在聚酮化合物和非核糖体肽代谢物的生物合成机制中可能比最初认识的更为普遍。该项目将集中在预测发生在蓝藻非核糖体肽生物合成途径中的HDD/SLiM相互作用的表征。该项目的目的是通过在E.大肠杆菌，并使用合成的底物类似物，以检查其酶活性。我们还将尝试使用X射线晶体学和NMR光谱学获得蛋白质的高分辨率结构信息。一旦表征，该系统将被利用来了解HDD/SLiM系统作为生物合成途径工程工具的潜力，包括使用来自不同代谢物的生物合成的组分生产混合装配线。