In planta chemical cross-linking and mass spectrometry analysis of protein structure and interaction in Arabidopsis

Xinliang Zhu, Fengchao Yu, Zhu YANG, Shichang Liu, Chen Dai, Xiaoyun Lu, Chenyu Liu, Weichuan Yu, Ning Li*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

Site-specific chemical cross-linking in combination with mass spectrometry analysis has emerged as a powerful proteomic approach for studying the three-dimensional structure of protein complexes and in mapping protein–protein interactions (PPIs). Building on the success of MS analysis of in vitro cross-linked proteins, which has been widely used to investigate specific interactions of bait proteins and their targets in various organisms, we report a workflow for in vivo chemical cross-linking and MS analysis in a multicellular eukaryote. This approach optimizes the in vivo protein cross-linking conditions in Arabidopsis thaliana, establishes a MudPIT procedure for the enrichment of cross-linked peptides, and develops an integrated software program, exhaustive cross-linked peptides identification tool (ECL), to identify the MS spectra of in planta chemical cross-linked peptides. In total, two pairs of in vivo cross-linked peptides of high confidence have been identified from two independent biological replicates. This work demarks the beginning of an alternative proteomic approach in the study of in vivo protein tertiary structure and PPIs in multicellular eukaryotes.

Original languageEnglish
Pages (from-to)1915-1927
Number of pages13
JournalProteomics
Volume16
Issue number13
DOIs
Publication statusPublished - 1 Jul 2016

Scopus Subject Areas

  • Biochemistry
  • Molecular Biology

User-Defined Keywords

  • Arabidopsis
  • Exhaustive cross-linked peptides identification tool
  • In planta chemical cross-linking
  • Interactomics
  • Mass spectrometry
  • Plant proteomics

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