Differential analysis of n-glycopeptide abundance and n-glycosylation site occupancy for studying protein n-glycosylation dysregulation in human disease

• Type of Material: Article
• Authors: Qi Zhang, Emory University; Cheng Ma, University of Texas Health Science Center at Houston; Lian Li, Emory University; Lih-Shen Chin, Emory University
• Language: English
• Date: 2021-06-20
• Publisher: Bio-protocol LLC
• Publication Version: Final Published Version
• Copyright Statement: © 2021 The Authors; exclusive licensee Bio-protocol LLC.
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.21769/BioProtoc.4059
• Title of Journal or Parent Work: Bio-protocol
• Volume: 11
• Issue: 12
• Start Page: e4059
• End Page: e4059
• Grant/Funding Information: This work was supported by the National Institutes of Health (NIH) Grants RF1AG057965 (to L.L.) and R56 AG059714 (to L.S.C.).; The Emory Center for Neurodegenerative Disease Brain Bank was supported in part by NIH Grants P50 AG025688 and P30 NS055077.
• Keywords: In vivo N-glycosylation sites; Alzheimer's disease; Mass spectrometry; N-glycosylation site occupancy; Protein N-glycosylation; Integrative glycoproteomics and proteomics; N-glycoproteomics; N-glycoproteome profiling
• Research Categories: Health Sciences, Pharmacology

Abstract

Protein N-glycosylation plays a vital role in diverse cellular processes, and dysregulated N-glycosylation is implicated in a variety of human diseases including neurodegenerative disorders and cancer. With recent advances in high-resolution mass spectrometry-based glycoproteomics technologies enabling large-scale N-glycoproteome profiling of disease and control samples, analysis of the large datasets has become a challenge. Here, we provide a protocol for the systems-level analysis of in vivo N-glycosylation sites on N-glycosylated proteins and their changes in human disease, such as Alzheimer's disease. The protocol includes quantitation and differential analysis of N-glycopeptide abundance, in addition to integrative N-glycoproteome and proteome data analyses, to determine disease-associated changes in N-glycosylation site occupancy and identify differentially N-glycosylated proteins in human disease versus control samples. This protocol can be modified and applied to study proteome-wide N-glycosylation alterations in response to different cellular stresses or pathophysiological states in other organisms or model systems.