How conformation matters: Ionic liquids as reaction media for oxidative folding and native chemical ligation of cysteine-containing peptides

考虑到离子液体在蛋白质和多肽化学中应用的重要性，表征离子对多肽结构和以多肽为反应物的反应的影响也是至关重要的。本项目主要研究含半胱氨酸多肽的两个反应：氧化折叠反应和天然化学连接反应。在初步研究中，从海洋锥螺中提取的结构紧凑的多个二硫键桥联神经毒素被证明在生物相容的离子液体中有效和正确地折叠成其生物活性构象。[C2mim][OAc]，[C2mim][Ots])。ILS对预制二级结构的稳定作用被认为是造成这一现象的原因。为了得到科学的解释，短小和中等大小的模型多肽以及天然的富含半胱氨酸的多肽将受到离子液体介导的氧化折叠。通过光谱测量(例如CD和荧光光谱)进行监测，将为多肽在离子液体中的嵌入提供深入的了解。将改变反应条件，以建立单个多肽折叠过程的优化，并探索是否有不同的参数影响多肽的稳定性和二硫键的形成。作为涉及半胱氨酸残基的第二种反应类型，NCL也将在离子液体中进行研究。在这里，对起始物质和反应产物(小蛋白质、蛋白质结构域)的稳定和增溶作用决定了合成结果。因此，该项目将提供新的方法，以获得规模化的含有半胱氨酸的多肽和蛋白质，从而能够对具有治疗潜力的这种复杂分子进行生物、生化和电生理研究。

Concerning the increased significance of ionic liquids for applications in protein and peptide chemistry it is as well essential to characterize the ion effects on peptide structures and reactions involving peptides as reactants. This project is focused on studies of two reactions involving cysteine-containing peptides, oxidative folding and native chemical ligation (NCL). In preliminary studies structurally compact multiple disulfide-bridged neurotoxins derived from marine cone snails were shown to efficiently and correctly fold into their biologically active conformation in biocompatible ionic liquids (e.g. [C2mim][OAc], [C2mim][OTs]). Stabilization of pre-formed secondary structures by ILs was suggested to be the cause of this observation. In order to derive a scientific explanation, short and medium-sized model peptides as well as native Cys-rich peptides will be subjected to ionic liquid-mediated oxidative folding. Monitoring by spectroscopic measurements (e.g. CD, and fluorescence spectroscopy) will provide insights into embedment of the peptides in ionic liquids. Reaction conditions will be varied to establish optimization of individual peptide folding processes and to explore whether there are distinct parameter influencing peptide stabilization and disulfide bridge formation. As a second reaction type involving cysteine residues NCL will be investigated in ionic liquids, too. Here, stabilizing and solubilising effects on both, the starting material as well as the reaction products (small proteins, protein domains) is determining the synthetic outcome. The project will therefore provide new methodologies to obtain cysteinecontaining peptides and proteins in a scale enabling biological, biochemical and electrophysiological studies for such complex molecules carrying therapeutic potential.