Fig. 3: Inhibition of glycan binding to RBD by MeαNeu5Ac and HS oligosaccharides.

a, Scatter plot of the relative change in normalized abundances (to 71) of released neutral and acidic glycans in the presence and absence of MeαNeu5Ac. Measurements were performed in aqueous ammonium acetate solutions (100 mM, pH 6.9, 25 °C) of RBD (10 μM) and each of the defined libraries (50 nM of each glycan) with 0, 200, and 500 µM MeαNeu5Ac. Circles and diamonds represent neutral and acidic glycans, respectively, and solid lines and dashed lines represent the median values and first and third quartiles, respectively. Statistical significance was calculated based on an unpaired Mann–Whitney test because the distribution of data failed numerous normality tests (D’Agostino–Pearson omnibus, Shapiro–Wilk and Kolmogorov–Smirnov); n = 3 independent experiments for each glycan. b, Change in normalized abundance of released glycan 69 following the addition of glycan 41 or MeαNeu5Ac to the solution. Measurements were performed in aqueous ammonium acetate solutions (100 mM, pH 6.9, 25 °C) of RBD (5 μM) and 69 (50 nM) in the absence and presence of 41 (triangle) or MeαNeu5Ac (circle) at concentrations ranging from 0.1 to 100 μM (41) or 500 μM (MeαNeu5Ac). Ions in the range of m/z 3,000–4,000 were subjected to HCD using a collision energy of 50 V. Data represent mean ± s.d.; n = 3 independent experiments. c, Zero-charge mass spectra of RBD (5 μM) alone and in the presence of glycan 69 (25 μM) or glycan 41 (50 μM) or a mixture of 69 (25 μM) and 41 (50 μM). An asterisk indicates a sodium adduct. Experiments were performed in aqueous ammonium acetate solutions (100 mM, pH 6.9, 25 °C). Deconvolution was performed using the Thermo BioPharma Finder Software.

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