摘要
C端用香豆素修饰的泛素分子(Ub-AMC)是研究蛋白质泛素化过程的重要探针.该探针分子的制备目前主要依靠生物表达结合化学修饰的方法,合成效率较低.本文使用多肽酰肼连接反应,发展出化学全合成Ub-AMC分子的新路线.该方法通过N到C顺序两次连接实现了目标分子的组装,再通过自由基脱硫反应得到天然结构的Ub-AMC分子,有望实现较大量的合成.通过酶学活性实验,证实了通过新方法合成的Ub-AMC具有预期的生物活性.
The C-terminal conjugate of ubiquitin with 7-amino-4-methylcoumarin (Ub-AMC) is an important probe for fluorescence-based analysis of deubiquitinating enzyme (DUB) activity. It is important to develop more efficient methods for the preparation of Ub-AMC because the currently available technology is still expensive for scaled-up production. In the present work, we report an efficient strategy for total chemical synthesis of Ub-AMC through ligation of peptide hydrazides. Three peptide segments are assembled via N-to-C sequential ligation and the resulting product is converted to Ub-AMC via TCEP-mediated desulfurization. The synthetic Ub-AMC is shown to have expected biological functions through the measurement of the DUB activity.
出处
《中国科学:化学》
CAS
CSCD
北大核心
2013年第8期1051-1058,共8页
SCIENTIA SINICA Chimica
基金
国家重点基础研究发展计划(973计划)(2013CB932800)
国家自然科学基金(31100524,31170817,20972148)支持
关键词
酰肼连接反应
多片段连接
去泛素化酶
泛素
ligation of peptide hydrazides
peptide segment condensation
deubiquitinating enzymes
ubiquitin