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Analysis of mutation-originated gain-of-glycosylation using mass spectrometry-based N-glycoproteomics
  • ZHIXIN TIAN,
  • Hailun Yang
ZHIXIN TIAN
Tongji University

Corresponding Author:[email protected]

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Hailun Yang
Tongji University
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Abstract

RATIONALE: A general N-glycoproteomics analysis pipeline has been established for characterization of mutation-related gain-of-glycosylation (GoG) at intact N-glycopeptide molecular level, generating comprehensive site and structure information of N-glycosylation. METHODS: This study focus on mutation-originated gain-of-glycosylation using mass spectrometry-based N-glycoproteomics analysis workflow. In brief, GoG intact N-glycopeptide databases were built, consisting of 2,701 proteins (potential GoG N-glycosites and amino acids derived from MUTAGEN, VARIANT and VAR_SEQ in UniProt) and 6,709 human N-glycans (≤50 sequence isomers per monosaccharide composition). We employed the site- and structure-specific N-glycoproteomics workflow utilizing intact N-glycopeptides search engine GPSeeker to identify GoG intact N-glycopeptides from parental breast cancer stem cells (MCF-7 CSCs) and adriamycin-resistant breast cancer stem cells (MCF-7/ADR CSCs). RESULTS: With the criteria of spectrum-level FDR control of ≤1%, we identified 88 and 96 GoG intact N-glycopeptides corresponding to 38 and 36 intact N-glycoproteins from MCF-7 CSCs and MCF-7/ADR CSCs, respectively. Among KEGG annotation of GoG N-glycoproteins, DNA polymerase eta (POLH), serine-protein kinase ATM (ATM) and cellular tumor antigen p53 (P53) were enriched in platinum drug resistance signal pathway. ATM, P53 and G2 and S phase-expressed protein 1 (GTSE1) were associated with p53 signaling pathway. CONCLUSIONS: The integration of site- and structure-specific N-glycoproteomics approach, conjugating with GoG characterization, provides a universal workflow for revealing comprehensive N-glycosite and N-glycan structure information of gain-of-glycosylation. The analysis of mutation-originated gain-of glycosylation can be extended GoG characterization to all the other N-glycoproteome systems including complex clinical tissues and body fluids.
21 Oct 2023Submitted to Rapid Communications in Mass Spectrometry
21 Oct 2023Assigned to Editor
21 Oct 2023Submission Checks Completed
21 Oct 2023Review(s) Completed, Editorial Evaluation Pending
05 Nov 2023Reviewer(s) Assigned
11 Feb 2024Review(s) Completed, Editorial Evaluation Pending
11 Feb 2024Editorial Decision: Revise Minor