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. 2022 Mar 29;11:e75545. doi: 10.7554/eLife.75545

Figure 7. Quantitative characterization of the disaccharide contents of heparan sulfate chains derived from Glypican-1 (GPC1), GPC5, and SDC4.

The recombinant forms of GPC1, GPC5, and SDC4 described in Figure 6 and Figure 6—figure supplement 1 were treated with a mixture of the heparinases I + II and III to release the disaccharide units of their heparan sulfate chains. These were then identified by their retention times in a analytical anion exchange HPLC setup using synthetic disaccharides as standards (Figure 7—figure supplement 1). For details, see Materials and methods.

(A) Representative elution profile of the disaccharide units derived from the heparan sulfate chains of GPC1 (black), GPC5 (blue), and SDC4 (red). (B) Statistical analysis of four independent experiments providing the relative abundances of heparan sulfate disaccharide units corresponding to the 12 standards (Figure 7—figure supplement 1) contained in GPC1 (black), GPC5 (blue), and SDC4 (red). Standard deviations are shown. Statistics were based on a two-way ANOVA test combined with a Bonferroni post-test (*, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001, and ****, p ≤ 0.0001).

Figure 7—source data 1. Raw data of the analytical HPLC experiments quantifying disaccharide units of the heparan sulfate chains of the proteoglycans indicated.

Figure 7.

Figure 7—figure supplement 1. Characterization of synthetic dissacharide standards corresponding to the building blocks of heparan sulfate chains using an analytical HPLC analysis.

Figure 7—figure supplement 1.

The standards used were commercial products (Iduron, UK). They were characterized for their retention times on a strong anion exchange column operated by a HPLC system. For experimental details, see Materials and methods.