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Promoting Tag Removal of a MBP-Fused Integral Membrane Protein by TEV Protease

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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.

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Acknowledgments

This work is supported by grants from the Ministry of Science and Technology (2016YFA0501200), the National Natural Science Foundation of China (31500603 and 21425523), and the Fundamental Research Funds for the Central Universities (WUT 2016IB006).

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Authors and Affiliations

  1. 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

    Yanke Chen & Jun Yang

  2. School of Chemistry, Chemical Engineering and Life Sciences, Wuhan University of Technology, Wuhan, 430070, China

    Qichang Li & Hao Xie

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  1. Yanke Chen
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  2. Qichang Li
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  3. Jun Yang
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  4. Hao Xie
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Correspondence to Jun Yang or Hao Xie.

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Chen, Y., Li, Q., Yang, J. et al. Promoting Tag Removal of a MBP-Fused Integral Membrane Protein by TEV Protease. Appl Biochem Biotechnol 181, 939–947 (2017). https://doi.org/10.1007/s12010-016-2260-z

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  • DOI: https://doi.org/10.1007/s12010-016-2260-z

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