title: Promoting Tag Removal of a MBP-Fused Integral Membrane Protein by TEV Protease.
abstract: Tag removal is a prerequisite issue for structural and functional analysis of affinity-purified membrane proteins. The present study took a MBP-fused membrane protein, MrpF, as a model to investigate the tag removal by TEV protease. Influences of the linking sequence between TEV cleavage site and MrpF on protein expression and predicted secondary structure were investigated. The steric accessibility of TEV protease to cleavage site of MBP-fused MrpF was explored. It was found that reducing the size of hydrophilic group of detergents and/or extending the linking sequence between cleavage site and target protein can significantly improve the accessibility of the cleavage site and promote tag removal by TEV protease.
authors.affiliation: Key Laboratory of Magnetic Resonance in Biological Systems, State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, Wuhan Center for Magnetic Resonance, Wuhan Institute of Physics and Mathematics, Chinese Academy of Sciences, Wuhan, 430071, China.
authors.forename: Yanke
authors.initials: Y
authors.lastname: Chen
authors.affiliation: School of Chemistry, Chemical Engineering and Life Sciences, Wuhan University of Technology, Wuhan, 430070, China.
authors.forename: Qichang
authors.initials: Q
authors.lastname: Li
authors.affiliation: Key Laboratory of Magnetic Resonance in Biological Systems, State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, Wuhan Center for Magnetic Resonance, Wuhan Institute of Physics and Mathematics, Chinese Academy of Sciences, Wuhan, 430071, China. yangjun@wipm.ac.cn.
authors.forename: Jun
authors.initials: J
authors.lastname: Yang
authors.affiliation: School of Chemistry, Chemical Engineering and Life Sciences, Wuhan University of Technology, Wuhan, 430070, China. h.xie@whut.edu.cn.
authors.forename: Hao
authors.initials: H
authors.lastname: Xie
mesh_terms: D001412:Bacillus subtilis / Q000737:chemistry*; D010450:Endopeptidases / Q000737:chemistry*; D058619:Maltose-Binding Proteins / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry*
chemical_list: D058619:Maltose-Binding Proteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D010450:Endopeptidases; C099488:TEV protease
keywords: Expression and purification; Integral membrane protein; Maltose binding protein; Steric occlusions; TEV protease; Tag removal