Expedient Total Synthesis of Small to Medium-Sized Membrane Proteins via Fmoc Chemistry

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• 作者：Ji-Shen Zheng ; Mu Yu ; Yun-Kun Qi ; Shan Tang ; Fei Shen ; Zhi-Peng Wang ; Liang Xiao ; Longhua Zhang ; Chang-Lin Tian ; Lei Liu
• 刊名：Journal of the American Chemical Society
• 出版年：2014
• 出版时间：March 5, 2014
• 年：2014
• 卷：136
• 期：9
• 页码：3695-3704
• 全文大小：682K
• 年卷期：v.136,no.9(March 5, 2014)
• ISSN：1520-5126

文摘

Total chemical synthesis provides a unique approach for the access to uncontaminated, monodisperse, and more importantly, post-translationally modified membrane proteins. In the present study we report a practical procedure for expedient and cost-effective synthesis of small to medium-sized membrane proteins in multimilligram scale through the use of automated Fmoc chemistry. The key finding of our study is that after the attachment of a removable arginine-tagged backbone modification group, the membrane protein segments behave almost the same as ordinary water-soluble peptides in terms of Fmoc solid-phase synthesis, ligation, purification, and mass spectrometry characterization. The efficiency and practicality of the new method is demonstrated by the successful preparation of Ser64-phosphorylated M2 proton channel from influenza A virus and the membrane-embedded domain of an inward rectifier K⁺ channel protein Kir5.1. Functional characterizations of these chemically synthesized membrane proteins indicate that they provide useful and otherwise-difficult-to-access materials for biochemistry and biophysics studies.