Profiling Glycoproteins Enriched by Multinanoparticle Protein Corona.

Area of Science:

• Biochemistry
• Proteomics
• Glycomics

Background:

• Biofluid analysis faces challenges due to complex protein composition and wide dynamic range.
• Nanoparticle (NP)-based enrichment methods show promise for improving proteomic sampling depth.
• The impact of NP enrichment on glycoproteome characterization is underexplored.

Purpose of the Study:

• To investigate nanoparticle (NP) enrichment with and without mixed-mode anion exchange (MAX) for glycopeptide characterization in human plasma and CSF.
• To assess the impact of NP enrichment on N- and O-linked glycoproteome coverage.
• To explore the application of NP enrichment for CSF analysis.

Main Methods:

• Utilized an automated commercial platform for NP enrichment using multinanoparticle protein corona.
• Applied subsequent MAX enrichment to a subset of samples.
• Analyzed N- and O-linked glycopeptides in human plasma and CSF.

Main Results:

• NP enrichment significantly enhanced N-glycoproteome coverage, especially in plasma, and improved detection of low-abundance N-glycopeptides.
• Combined NP and MAX enrichment further boosted N-glycoproteome improvements.
• NP enrichment alone was superior for O-glycoproteome analysis, suggesting MAX preferentially enriches N-glycopeptides.
• First-time application of NP enrichment to CSF revealed increased proteome and glycoproteome depth, including extracellular vesicle biomarkers.

Conclusions:

• NP enrichment, particularly when combined with MAX, is effective for enhancing N-glycoproteome coverage in human biofluids.
• NP enrichment alone is optimal for O-glycoproteome analysis.
• NP enrichment offers a valuable framework for glycoprotein analysis in complex biofluids like CSF, aiding biomarker discovery.