Chemo-enzymatic platform for the discovery of natural and functionalized sactipeptides

多肽是细菌起源的小蛋白质家族的成员。它们的编码序列被组织在基因簇中，这些基因簇编码这些核糖体合成的微小蛋白合成后修饰所需的酶。它们的标志是至少存在一个分子内硫醚桥，该桥是由半胱氨酸硫醇与伙伴氨基酸的α碳原子连接而形成的。通过这种非常不寻常的连接，只有几十个氨基酸长度的生物分子被形成，表现出非凡的化学稳定性和构象刚性。多肽通常具有抗菌活性，因此是用作抗生素的潜在候选者。通过数据库挖掘，已经在细菌基因组中发现了许多假定的多肽。只有五种多肽的硫醚键的细节是已知的。该项目的主要目标是通过结合分子生物学、生物化学和肽化学的方法，对迄今尚未开发的sactipeptide进行深入的表征。这些知识将用于使用蛋白质设计和定向蛋白质进化策略生成具有新功能的多肽。迄今为止，对疾病相关靶分子具有定制亲和力的多肽尚未被描述。它们成功生成的先决条件是对各自的皂素合成酶对皂素前体蛋白的分子识别有详细的了解，皂素合成酶是化学酶合成非天然皂素肽的工具。因此，硫醚形成酶识别多肽前体的序列和结构参数也将在本项目的框架内进行研究。

Sactipeptides are members of a family of small proteins of bacterial origin. Their coding sequence is organized in gene clusters that encode enzymes required for the postsynthetic modification of these ribosomally synthesized miniproteins. Their hallmark is the presence of at least one intramolecular thioether bridge that is formed by the linkage of a cysteine thiol with the alpha carbon atom of a partner amino acid. Through this very uncommon linkage, biomolecules with only a few dozens of amino acids in length are formed that display extraordinary chemical stability and conformational rigidity. Sactipeptides often show antibacterial activity and therefore are potential candidates for their use as antibiotics. Numerous putative sactipeptides have been identified in bacterial genomes through database mining. Only for five sactipeptides the details of their thioether linkages are known. The main goal of the proposed project is the in depth characterization of up to now unexplored sactipeptides by application of a combination of methods of molecular biology, biochemistry, and peptide chemistry. This knowledge will be used for the generation of sactipeptides with novel functionalities using protein design and directed protein evolution strategies. Sactipeptides with tailor-made affinities towards disease-related target molecules have not been described to date. A prerequisite of their successful generation is a detailed knowledge about the molecular recognition of sactipeptide precursor proteins by the respective sactionine synthase enzyme that serves as tool for the chemo-enzymatic synthesis of non-natural sactipeptides. As a consequence, sequence and structure parameters of sactipeptide precursor recognition by the thioether forming enzymes will also be investigated in the frame of this project.