Catalyst-free "click" reactions for the direct conjugation of metal complexes to bio(macro)molecules

由于这些化合物中存在大量的活性基团，因此对含有非天然官能团的多肽和蛋白质等生物大分子进行温和的位置选择性修饰是一个巨大的挑战。与此同时，这种混合系统作为研究基本生物过程的工具提供了巨大的前景，它们也可能在医学诊断和治疗中发现有趣的应用。生物正交“点击反应”，如铜催化的1，3-偶极叠氮-炔环加成反应(CuAAC)，目前被广泛用于为生物大分子引入新的官能团。然而，由于需要一种潜在的有毒过渡金属催化剂，阻碍了CuAAC和其他偶联方法在生命系统中的应用，因此，本项目的目标是开发新型温和的、无催化剂的功能金属络合物与多肽和蛋白质的偶联方法。然而，与其他最近的生物正交偶联反应不同，如菌株促进的叠氮化物-炔环加成反应，我们的工作将专门基于金属固有的反应性。我们战略的核心将是取代惰性金属叠氮化物与炔功能化的生物大分子在“iClick反应”中的直接反应，形成金属-三氮唑键。金属中心和配体球体的灵活变化将使我们能够调整偶联伙伴的电子和空间性质，以确保快速反应动力学，并引入各种功能来探索生物系统。然后，通过这种方法制备的杂化金属络合物-多肽生物结合物将在氨基酸、核苷酸和生物硫醇等常见细胞成分的挑战实验中研究三氮唑键的稳定性。虽然最初的工作将集中在简单模型多肽的使用上，但随后它们的复杂性将增加到具有潜在生物相关性的较大载体多肽。选定的金属络合物-多肽杂交物将被测试它们对人类癌细胞的生物活性。在建立了一组金属反应伙伴和手头的耦合条件后，该项目将采取初步步骤，利用“iClick”反应性对功能蛋白质进行修饰，为这些系统引入新的基于金属的功能。

The mild and site-selective modification of biomacromolecules like peptides and proteins with non-natural functional groups is a significant challenge due to the multitude of reactive groups these compounds present. At the same time, such hybrid systems offer great promise as tools to study fundamental biological processes and they might also find interesting applications in medical diagnosis and therapy. Bioorthogonal "click reactions" like the copper-catalyzed 1,3-dipolar azide-alkyne cycloaddition (CuAAC) are now heavily utilized to introduce new functionalities to biomacromolecules. However, the application of the CuAAC and other coupling methods in living systems is hampered by the need for a potentially toxic transition metal catalyst.Therefore, it is the aim of the present project to develop novel mild and catalyst-free methods for the conjugation of functional metal complexes to peptides and proteins. However, in contrast to other very recent bioorthogonal coupling reactions like the strain-promoted azide-alkyne cycloaddition, our work will be specifically based on a metal-inherent reactivity. At the core of our strategy will be the direct reaction of substitutionally inert metal azides with alkyne-functionalized biomacromolecules in an "iClick reaction" to form a metal-triazolate linkage. Facile variation of the metal center and coligand sphere will allow us to tune the electronic and steric properties of the coupling partners to ensure fast reaction kinetics and introduce a wide variety of functionalities to probe biological systems.The hybrid metal complex-peptide bioconjugates prepared this way will then be studied for stability of the triazolate linkage in challenge experiments with common cellular constituents like amino acids, nucleotides, and biogenic thiols. While initial work will focus on the use of simple model peptides, their complexity will then subsequently be increased to larger carrier peptides of potential biological relevancy. Selected metal complex-peptide hybrids will be tested for their biological activity on human cancer cells.With an established set of metal reaction partners and coupling conditions at hand, this project will then take initials steps towards the modification of functional proteins using the "iClick" reactivity to introduce novel, metal-based functionality to these systems.