N Glycan profiling in plasma proteins of semi supercentenarians using sialic acid linkage specific derivatization and negative ion MALDI TOF MS

Significance of the Topic

N-glycosylation of plasma proteins and the specific linkages of sialic acids play vital roles in modulating immune responses, inflammation, and overall homeostasis. Detailed profiling of these glycan structures in long-lived individuals can reveal biomarkers and mechanisms associated with healthy aging and extreme longevity.

Objectives and Study Overview

This study aimed to compare plasma N-glycan profiles of semi-supercentenarians (SSCs, ≥105 years old) against three younger control groups (70s, 80s, 90s). By applying sialic acid linkage-specific derivatization followed by negative-ion MALDI-TOF mass spectrometry and multivariate analysis, the investigation sought to identify age-dependent glycan changes linked to extreme lifespan.

Methodology and Used Instrumentation

Plasma samples were pooled (4–5 individuals per pool) and depleted of albumin and IgG. N-glycans were released enzymatically with PNGase F and captured with hydrazide beads (BlotGlyco®). Sialic acids were stabilized via linkage-specific alkylamidation (SALSA) using the SialoCapper™-ID Kit, with 2-AA labeling to improve ionization.

• Mass spectrometer: Shimadzu MALDI-8030 in negative-ion linear mode.
• Matrix: super-DHB.
• Data acquisition: eMSTAT solution for automated plate measurement, lock mass calibration at m/z 2426.03.
• Data processing: peak picking with 0.1% intensity threshold, 3 Da tolerance, Pareto scaling, PLS-DA multivariate analysis.

Main Results and Discussion

Multivariate PLS-DA separated SSCs from younger groups, highlighting specific N-glycan alterations with age. A tri-sialylated glycan carrying one α2,3-linked and two α2,6-linked sialic acids was markedly elevated in SSCs. General trends included incremental increases of highly sialylated structures and shifts in fucosylated species, suggesting adaptations in anti-inflammatory glycoforms over the human lifespan.

Benefits and Practical Applications

Linkage-specific glycan profiling offers:

• Potential biomarkers for healthy aging and longevity.
• Tools to monitor chronic inflammation and immune status in elderly populations.
• High-throughput screening capability for clinical or epidemiological studies.

Future Trends and Potential Applications

Emerging developments may include microfluidic integration for reduced sample consumption, MS imaging of glycosylation patterns in tissues, single-cell glycomics, and integration with proteomics and transcriptomics for systems-level insights into aging.

Conclusion

By combining sialic acid linkage-specific derivatization with negative-ion MALDI-TOF MS, the study successfully identified age-related plasma glycan signatures associated with extreme human longevity. These findings pave the way for novel biomarkers and a deeper understanding of glycosylation’s role in healthy aging.

References

1. Miura Y et al. PLoS ONE. 2015;10:e0142645.
2. Miura Y et al. Biochim Biophys Acta Gen Subj. 2018;1862:1462–1471.
3. Nishikaze A et al. Anal Chem. 2017;89:2353–2360.