Functional Hybrid Natural Product Synthases by Tracking Acyl Carrier Protein Binding and Conformational Dynamics

项目总结 微生物产生结构不同的分子，其中许多已经被成功地改变了用途 人类作为药剂。这些分子是由多酶组装而成的，这是 使用酰基载体蛋白(ACP)修饰和转移化学中间体。自然界的理性再设计 酶组件为生产新抗生素提供了一条令人兴奋的可能途径，但任何 重新设计方法取决于对导致化学生产的杂交ACP的彻底理解- 酶的相互作用。这项研究的目标是了解ACP如何与它们的两个分子相互作用 货物和辅助酶。ACP的动态结构及其蛋白质的瞬变性质 相互作用使得这类蛋白质的研究特别具有挑战性。因此，ACP的相互作用将是 使用现场特定振动光谱和沉积速度实验观察到的创新 对快速构象变化和弱的蛋白质-蛋白质相互作用敏感的方法。信息 这项工作将在从传统方法获得的数据的背景下进行分析，以确定 候选杂交ACP-酮合成酶配对能够产生新结构的分子。的能力 将对产生聚酮的杂交对进行评估。这些研究的结果将指导未来 具有潜在药用活性的新型小分子的生物合成。

PROJECT SUMMARY Microorganisms produce structurally diverse molecules, many of which have been successfully repurposed by mankind as pharmaceutical agents. These molecules are manufactured by multi-enzyme assemblies, which use acyl carrier proteins (ACPs) to modify and transfer chemical intermediates. Rational redesign of natural enzyme assemblies presents an exciting possible route to produce new antibiotics, but the success of any redesign approach depends on a thorough understanding of what leads to chemically productive hybrid ACP- enzyme interactions. The goal of this study is to understand how ACPs interact with both their molecular cargoes and partner enzymes. The dynamic structures of ACPs and the transient nature of their protein interactions make this class of proteins particularly challenging to study. Therefore, ACP interactions will be observed using site-specific vibrational spectroscopy and sedimentation velocity experiments, an innovative approach that is sensitive to fast conformational changes and weak protein-protein interactions. Information from this work will be analyzed in the context of data acquired from traditional methodologies to identify candidate hybrid ACP-ketosynthase partners capable of producing molecules of novel structure. The ability of the hybrid pairs to produce polyketides will be evaluated. Results from these studies will guide the future biosynthesis of novel small molecules with potential pharmaceutical activity.