Relative Quantification of N‑Glycopeptide Sialic Acid Linkage Isomers by Ion Mobility Mass Spectrometry

Sialic
acids decorate the surface of glycoproteins and play important
roles in a variety of pathological processes. Although the mass spectrometry
(MS) based linkage-specific analysis of sialylated N-glycopeptide
is developing rapidly, quantitative analysis of these isomers still
remains a challenge. Herein, we reported a novel quantitative strategy
that can unambiguously identify and relatively quantify linkage-specific
N-glycopeptides using ion mobility mass spectrometry (IM-MS). Without
the assistance of derivatization, this method can relatively quantify
sialic acid isomers of intact glycopeptides by using their characteristic
fragment ions in IM-MS. Moreover, good linearity (R2 > 0.99) of relative quantification within a dynamic
range of 2 orders of magnitude and high reproducibility (coefficient
of variation (CV) < 10%, n = 3) were demonstrated.
Finally, our results illustrated the aberrant sialylation of haptoglobin
(Hp) in hepatocellular carcinoma (HCC), where the ratios of α2,3
to α2,6 sialylation of seven N-glycopeptides were found to be
significantly altered (p < 0.01) in HCC individuals
(n = 27) compared with healthy controls (n = 27).