title: Robust Chemical Synthesis of Membrane Proteins through a General Method of Removable Backbone Modification.
abstract: Chemical protein synthesis can provide access to proteins with post-translational modifications or site-specific labelings. Although this technology is finding increasing applications in the studies of water-soluble globular proteins, chemical synthesis of membrane proteins remains elusive. In this report, a general and robust removable backbone modification (RBM) method is developed for the chemical synthesis of membrane proteins. This method uses an activated O-to-N acyl transfer auxiliary to install in the Fmoc solid-phase peptide synthesis process a RBM group with switchable reactivity toward trifluoroacetic acid. The method can be applied to versatile membrane proteins because the RBM group can be placed at any primary amino acid. With RBM, the membrane proteins and their segments behave almost as if they were water-soluble peptides and can be easily handled in the process of ligation, purification, and mass characterizations. After the full-length protein is assembled, the RBM group can be readily removed by trifluoroacetic acid. The efficiency and usefulness of the new method has been demonstrated by the successful synthesis of a two-transmembrane-domain protein (HCV p7 ion channel) with site-specific isotopic labeling and a four-transmembrane-domain protein (multidrug resistance transporter EmrE). This method enables practical synthesis of small- to medium-sized membrane proteins or membrane protein domains for biochemical and biophysical studies.
authors.affiliation: High Magnetic Field Laboratory, Chinese Academy of Sciences, and School of Life Sciences, University of Science and Technology of China , Hefei 230031, China.
authors.forename: Ji-Shen
authors.initials: JS
authors.lastname: Zheng
authors.affiliation: High Magnetic Field Laboratory, Chinese Academy of Sciences, and School of Life Sciences, University of Science and Technology of China , Hefei 230031, China.
authors.forename: Yao
authors.initials: Y
authors.lastname: He
authors.affiliation: Tsinghua-Peking Center for Life Sciences, Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology (Ministry of Education), Department of Chemistry, Tsinghua University , Beijing 100084, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Zuo
authors.affiliation: High Magnetic Field Laboratory, Chinese Academy of Sciences, and School of Life Sciences, University of Science and Technology of China , Hefei 230031, China.
authors.forename: Xiao-Ying
authors.initials: XY
authors.lastname: Cai
authors.affiliation: Tsinghua-Peking Center for Life Sciences, Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology (Ministry of Education), Department of Chemistry, Tsinghua University , Beijing 100084, China.
authors.forename: Shan
authors.initials: S
authors.lastname: Tang
authors.affiliation: Tsinghua-Peking Center for Life Sciences, Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology (Ministry of Education), Department of Chemistry, Tsinghua University , Beijing 100084, China.
authors.forename: Zhipeng A
authors.initials: ZA
authors.lastname: Wang
authors.affiliation: High Magnetic Field Laboratory, Chinese Academy of Sciences, and School of Life Sciences, University of Science and Technology of China , Hefei 230031, China.
authors.forename: Long-Hua
authors.initials: LH
authors.lastname: Zhang
authors.affiliation: High Magnetic Field Laboratory, Chinese Academy of Sciences, and School of Life Sciences, University of Science and Technology of China , Hefei 230031, China.
authors.forename: Chang-Lin
authors.initials: CL
authors.lastname: Tian
authors.affiliation: Tsinghua-Peking Center for Life Sciences, Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology (Ministry of Education), Department of Chemistry, Tsinghua University , Beijing 100084, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Liu
mesh_terms: D008565:Membrane Proteins / Q000138:chemical synthesis* / Q000737:chemistry; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry; D060327:Solid-Phase Synthesis Techniques / Q000379:methods*
chemical_list: D008565:Membrane Proteins; D010455:Peptides; C015462:polyarginine