Glassey Emerson, Zhang Zhengan, King Andrew M, Niquille David L, Voigt Christopher A
Synthetic Biology Center, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
Nat Chem. 2025 Feb;17(2):233-245. doi: 10.1038/s41557-024-01685-9. Epub 2025 Jan 7.
In nature, peptides are enzymatically modified to constrain their structure and introduce functional moieties. De novo peptide structures could be built by combining enzymes from different pathways, but determining the rules of their use is difficult. We present a biophysical model to combine enzymes sourced from bacterial ribosomally synthesized and post-translationally modified peptide (RiPP) gene clusters. Using a pipeline to evaluate more than 1,000 peptides, the model was parameterized under uniform conditions in Escherichia coli for enzymes from different classes (graspetide, spliceotide, pantocin, cyanobactin, glycocin, lasso peptide and lanthipeptide). Synthetic leader peptides with recognition sequences for up to three enzymes were designed to modify core sequences sharing no identity to natural RiPPs. Empirically, RiPPs with the desired modifications constituted 7-67% of the total peptides produced, and 6 of our 8 peptide designs were successfully modified. This work is an example of the design of enzyme-modified peptides and libraries, using a framework that can be expanded to include new enzymes and chemical moieties.
在自然界中，肽会通过酶促修饰来限制其结构并引入功能基团。可以通过组合来自不同途径的酶来构建从头合成的肽结构，但确定其使用规则却很困难。我们提出了一个生物物理模型，用于组合源自细菌核糖体合成及翻译后修饰肽（RiPP）基因簇的酶。使用一个管道来评估1000多种肽，该模型在大肠杆菌的统一条件下针对不同类别的酶（graspetide、spliceotide、pantocin、氰基细菌素、糖霉素、套索肽和羊毛硫肽）进行了参数化。设计了具有多达三种酶识别序列的合成前导肽，以修饰与天然RiPPs无同源性的核心序列。根据经验，具有所需修饰的RiPPs占产生的总肽的7%-67%，我们的8种肽设计中有6种成功得到了修饰。这项工作是酶修饰肽和文库设计的一个例子，使用的框架可以扩展以纳入新的酶和化学基团。
