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. 2019 Sep 13;165(11):1181–1197. doi: 10.1099/mic.0.000851

Fig. 5.

Fig. 5.

SiaPG cleaves α2-3 and α2-6 linked Neu5Ac from erythropoietin (EPO), though diacetylated sialic acid is retained. EPO was digested by SiaPG for 18 hours at 37 °C, and subjected to UHPLC-MS/MS. A deviation of MS2 ion m/z of±0.2 was permitted during spectral assignment. Sialylated structures may possess different isomers to those depicted (i.e. it is uncertain which terminal galactose residue(s) in the branched chains are sialylated). The top panel shows the UHPLC chromatogram for undigested EPO, the numbers indicate different EPO glycans, structures and further details of which are shown in Table S1. Lower panels show the UHPLC chromatogram for SiaPG digested EPO (orange), with three overlays showing MS1 ions which when fragmented produced MS2 ions with m/z corresponding to the precursor ion for Neu5Ac trisaccharide (GlcNAc-Gal-Neu5Ac, m/z=~657.25, green overlay),diactylated sialic acid trisaccharide (GlcNAc-Gal-Neu5,9Ac, m/z=~699.25, blue overlay, middle), and triacetylated sialic acid trisaccharide (GlcNAc-Gal-Neu5,8,9Ac, m/z=~741.25,brown overlay, lower).