Identification of Protein Direct Interactome with Genetic Code Expansion and Search Engine OpenUaa

Chao Liu, Ting Wu, Xin Shu, Shang-Tong Li, Daniel R. Wang, Nanxi Wang, Rong Zhou, Hao Yang, Hong Jiang, Ivo A. Hendriks, Pengyun Gong, Long Zhang, Michael L. Nielsen, Kui Li, Lei Wang, Bing Yang

Research output: Contribution to journalJournal articleResearchpeer-review

17 Citations (Scopus)

Abstract

Protein crosslinks occur endogenously such as modifications by ubiquitin-like proteins for signaling, or exogenously through genetically encoded chemical crosslinkers (GECX) for studying elusive protein-protein interactions. However, it remains challenging to identify these protein crosslinks efficiently at the proteomic scale. Herein, software OpenUaa is developed for identifying protein crosslinks generated by genetically encoded unnatural amino acids and endogenous protein conjugation. OpenUaa features inclusive and open search capability, dramatically improving identification sensitivity and coverage. Integrating GECX with OpenUaa, the direct interactome of thioredoxin is identified in Escherichia coli cells, yielding 289 crosslinked peptides and corresponding to 205 direct binding protein of thioredoxin. These identified direct binders provide evidence for thioredoxin's regulation of redox state and mitochondria energy metabolism. When identifying endogenous conjugation of small ubiquitin-like modifier (SUMO), OpenUaa also markedly improves coverage of SUMOylated peptides by ≈92%, revealing new SUMO targets. GECX-OpenUaa will enable efficient identification of direct interactomes of various proteins in live cells.

Original languageEnglish
Article number2000308
JournalAdvanced Biology
Volume5
Issue number3
Number of pages11
ISSN2701-0198
DOIs
Publication statusPublished - 2021

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