Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, 52074, Aachen, Germany.
DWI-Leibniz-Institute for Interactive Materials, Forckenbeckstraβe 50, 52074, Aachen, Germany.
Chemistry. 2020 Oct 27;26(60):13568-13572. doi: 10.1002/chem.202002740. Epub 2020 Aug 18.
Staphylococcus aureus sortase A (SaSrtA) is widely used for site-specific protein modifications, but it lacks the robustness for performing bioconjugation reactions at elevated temperatures or in presence of denaturing agents. Loop engineering and subsequent head-to-tail backbone cyclization of SaSrtA yielded the cyclized variant CyM6 that has a 7.5 °C increased melting temperature and up to 4.6-fold increased resistance towards denaturants when compared to the parent rM4. CyM6 gained up to 2.6-fold (vs. parent rM4) yield of conjugate in ligation of peptide and primary amine under denaturing conditions.
金黄色葡萄球菌天冬酰胺酰基内肽酶 A（SaSrtA）被广泛用于定点蛋白质修饰，但它缺乏在高温或变性剂存在下进行生物偶联反应的稳健性。SaSrtA 的环化变体 CyM6 通过环化工程和随后的从头至尾骨干环化得到，与亲本 rM4 相比，其熔点提高了 7.5°C，对变性剂的抗性提高了 4.6 倍。与亲本 rM4 相比，在变性条件下连接肽和伯胺时，CyM6 的偶联产物产率提高了 2.6 倍（vs. 亲本 rM4）。
