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Background <p>It is crucial to investigate protein functions in specific subcellular environments. Cross-linking mass spectrometry is a powerful tool to map the direct interactome of proteins by identifying inter-protein cross-links. However, it is challenging to identify inter-protein cross-linked peptides due to their low abundance.</p> Results <p>We chemically synthesize the cross-linkers ePDES1 and ePDES2 with an alkyne group and a compound with azide linked to a phosphate group to enrich for cross-linked peptides.</p> Conclusion <p>Based on the high-quality cross-linking spectra of ePDES1 and ePDES2, our methods achieve the identification of hundreds of directly interacting proteins or substrates of thioredoxins in the nucleus and mitochondria.</p>

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Enrichable cross-linkers for mapping direct protein interactions

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年7月15日

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  • Ting Wu,
  • Hang-Xu Zhou,
  • Jing Tian,
  • Rong Zhou,
  • Shangwei Huangfu,
  • Bi-Kun Jin,
  • Anna Sablina,
  • Fangfang Zhou,
  • Hongli Chen,
  • Shibing Tang,
  • Long Zhang,
  • Bing Yang

摘要

Background

It is crucial to investigate protein functions in specific subcellular environments. Cross-linking mass spectrometry is a powerful tool to map the direct interactome of proteins by identifying inter-protein cross-links. However, it is challenging to identify inter-protein cross-linked peptides due to their low abundance.

Results

We chemically synthesize the cross-linkers ePDES1 and ePDES2 with an alkyne group and a compound with azide linked to a phosphate group to enrich for cross-linked peptides.

Conclusion

Based on the high-quality cross-linking spectra of ePDES1 and ePDES2, our methods achieve the identification of hundreds of directly interacting proteins or substrates of thioredoxins in the nucleus and mitochondria.