ja500222u_si_001.pdf (2.52 MB)
Expedient Total Synthesis of Small to Medium-Sized Membrane Proteins via Fmoc Chemistry
journal contribution
posted on 2014-03-05, 00:00 authored by Ji-Shen Zheng, Mu Yu, Yun-Kun Qi, Shan Tang, Fei Shen, Zhi-Peng Wang, Liang Xiao, Longhua Zhang, Chang-Lin Tian, Lei LiuTotal
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.