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Covalent organic framework-coated magnetic graphene as a novel support for trypsin immobilization

Analytical and Bioanalytical Chemistry volume 409, pages 2179–2187 (2017)Cite this article

Abstract

Deep and efficient proteolysis is the critical premise in mass spectrometry-based bottom-up proteomics. It is difficult for traditional in-solution digestion to meet the requirement unless prolonged digestion time and enhanced enzyme dosage are employed, which makes the whole workflow time-consuming and costly. The abovementioned problems could be effectively ameliorated by anchoring many proteases on solid supports. In this work, covalent organic framework-coated magnetic graphene (MG@TpPa-1) was designed and prepared as a novel enzyme carrier for the covalent immobilization of trypsin with a high degree of loading (up to 268 μg mg−1). Profiting from the advantages of magnetic graphene and covalent organic frameworks, the novel trypsin bioreactor was successfully applied for the enzymatic digestion of a model protein with dramatically improved digestion efficiency, stability, and reusability. Complete digestion could be achieved in a time period as short as 2 min. For the digestion of proteins extracted from Amygdalus pedunculata, a total of 2833 protein groups were identified, which was slightly more than those obtained by 12 h of in-solution digestion (2739 protein groups). All of the results demonstrate that MG@TpPa-1-trypsin is an excellent candidate for sample preparation in a high-throughput proteomics analysis.

Covalent organic frameworks-coated magnetic graphene was prepared as novel carrier for highly efficient tryptic immobilization

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Acknowledgements

This work was supported by the National Natural Science Foundation of China (21675125, 21606181, 21275159, and 21235001), the National Key Program for Basic Research of China (2013CB911204 and 2016YFA0501403), the National Key Program for Scientific Instrument and Equipment Development (2012YQ12004407, 2011YQ06008408, and 2013YQ14040506). This work was also partly supported by the Amygdalus pedunculata Engineering Technology Research Center of State Forestry Administration and the Key Laboratory of Yulin Desert Plants Resources.

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Authors and Affiliations

  1. Key Laboratory of Synthetic and Natural Function Molecule Chemistry of Ministry of Education, College of Chemistry and Materials Science, Northwest University, Xi’an, Shaanxi, 710069, China

    Heping Wang & Yehua Shen

  2. State Key Laboratory of Proteomics, National Center for Protein Science Beijing, Beijing Institute of Radiation Medicine, Beijing, 102206, China

    Heping Wang, Fenglong Jiao, Fangyuan Gao, Xinyuan Zhao, Yan Zhao, Yangjun Zhang & Xiaohong Qian

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  1. Heping Wang
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  2. Fenglong Jiao
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  3. Fangyuan Gao
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  4. Xinyuan Zhao
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  5. Yan Zhao
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  6. Yehua Shen
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Correspondence to Yehua Shen, Yangjun Zhang or Xiaohong Qian.

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Wang, H., Jiao, F., Gao, F. et al. Covalent organic framework-coated magnetic graphene as a novel support for trypsin immobilization. Anal Bioanal Chem 409, 2179–2187 (2017). https://doi.org/10.1007/s00216-016-0163-z

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  • DOI: https://doi.org/10.1007/s00216-016-0163-z

Keywords

  • Covalent organic framework
  • Immobilized trypsin
  • Magnetic graphene