Design and Evolution of Enhanced Peptide-Peptide Ligation for Modular Transglutaminase Assembly.
Design and Evolution of Enhanced Peptide-Peptide Ligation for Modular Transglutaminase Assembly.
复制标题
DOI：
10.1021/acs.bioconjchem.3c00122
复制
发表时间：
2023-06-21
影响因子：
4.7
通讯作者：
Howarth, Mark
中科院分区：
文献类型：
--
作者：
Keeble, Anthony H.;Wood, Dominic P.;Howarth, Mark
需要强大而精确的工具来增强合成纳米结构的功能和弹性。在这里，我们采用定向进化和理性设计，从细菌粘附蛋白中构建了一种快速作用的分子超级胶水。我们已经生成了snopligase2偶联系统，这是一种在SnoopTag2和DogTag2肽之间有效转化的遗传编码途径。通过噬菌体展示筛选，选择每个肽段进行快速反应。优化后的试剂完成度超过99%，可与不同的缓冲液、pH值和温度兼容，可将反应速度提高1000倍以上。snopligase2在哺乳动物分泌途径中指导特定反应，允许在质膜上共价显示。转谷氨酰胺酶2 （TG2）在哺乳动物细胞表面和细胞外基质中具有相互作用和底物网络。我们表达了一个具有抗氧化失活和最小自反应性的修饰TG2。snopligase2使TG2通过转化生长因子α (TGFα)实现功能化，这是通过遗传融合无法实现的。TG2:TGFα偶联物保留转氨酶活性，稳定锚定TGFα在细胞外环境中的信号激活，并重新编程细胞行为。这种模块化工具箱应该为分子组装创造新的机会，无论是新的生物材料还是复杂的细胞环境。
Robust and precise tools are needed to enhance the functionality and resilience of synthetic nanoarchitectures. Here, we have employed directed evolution and rational design to build a fast-acting molecular superglue from a bacterial adhesion protein. We have generated the SnoopLigase2 coupling system, a genetically encoded route for efficient transamidation between SnoopTag2 and DogTag2 peptides. Each peptide was selected for rapid reaction by phage display screening. The optimized set allows more than 99% completion and is compatible with diverse buffers, pH values, and temperatures, accelerating the reaction over 1000-fold. SnoopLigase2 directs a specific reaction in the mammalian secretory pathway, allowing covalent display on the plasma membrane. Transglutaminase 2 (TG2) has a network of interactions and substrates amidst the mammalian cell surface and extracellular matrix. We expressed a modified TG2 with resistance to oxidative inactivation and minimal self-reactivity. SnoopLigase2 enables TG2 functionalization with transforming growth factor alpha (TGFα) in routes that would be impossible through genetic fusion. The TG2:TGFα conjugate retained transamidase activity, stably anchored TGFα for signal activation in the extracellular environment, and reprogrammed cell behavior. This modular toolbox should create new opportunities for molecular assembly, both for novel biomaterials and complex cellular environments.
登录
查看更多内容
影响因子：
5.6
作者：
Cobaugh, Christian W.;Almagro, Juan C.;Georgiou, George
通讯作者：
Georgiou, George
影响因子：
5.7
作者：
Izore, Thierry;Contreras-Martel, Carlos;Dessen, Andrea
通讯作者：
Dessen, Andrea
DOI：
10.1007/978-1-4939-2272-7_12
发表时间：
2015
期刊：
Methods in molecular biology (Clifton, N.J.)
影响因子：
--
作者：
Fairhead, Michael;Howarth, Mark
通讯作者：
Howarth, Mark
影响因子：
8.6
作者：
Keeble AH;Yadav VK;Ferla MP;Bauer CC;Chuntharpursat-Bon E;Huang J;Bon RS;Howarth M
通讯作者：
Howarth M
影响因子：
29.4
作者：
Jacucci, Gianni;Schertel, Lukas;Vignolini, Silvia
通讯作者：
Vignolini, Silvia