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The substrate specificity of proteases largely determines their function and biological role. Methods that can profile a protease’s specificity with high detail are valuable tools for protease research. Synthetic combinatorial peptide libraries offer a large array of potential substrates that can be tested in a single experiment. Here, we describe the design and synthesis of a combinatorial peptide library using the one-bead-one-compound approach, followed by treatment with a protease, i.e., Pro-Pro endopeptidase (PPEP), and subsequent LC-MS/MS and data analysis.

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Protease Specificity Profiling Using Synthetic Combinatorial Peptide Libraries and Mass Spectrometry

  • 实验方案

  • 发表日期：

    2025年

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  • Bart Claushuis,
  • Robert A. Cordfunke,
  • Arnoud H. de Ru,
  • Peter A. van Veelen,
  • Jeroen Corver,
  • Jan W. Drijfhout,
  • Paul J. Hensbergen

摘要

The substrate specificity of proteases largely determines their function and biological role. Methods that can profile a protease’s specificity with high detail are valuable tools for protease research. Synthetic combinatorial peptide libraries offer a large array of potential substrates that can be tested in a single experiment. Here, we describe the design and synthesis of a combinatorial peptide library using the one-bead-one-compound approach, followed by treatment with a protease, i.e., Pro-Pro endopeptidase (PPEP), and subsequent LC-MS/MS and data analysis.