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1.
Figure 2

Figure 2. Fold change plot of the abundances of Δ4,5-unsaturated disaccharides from control and HSulf2 treated bovine HS samples.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

Disaccharide abundances from Fig. 1A are shown as fold-change differences relative to control samples. Error bars are calculated from triplicate experiments and are reported as +/− S.D.

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.
2.
Figure 8

Figure 8. Effect of HSulf2 digestion on abundances of lyase-resistant tetrasaccharides in exhaustive digests of bovine HS.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

Two tetrasaccharides that resist exhaustive lyase digestion were quantified in control and HSulf2 treated bovine HS samples. The black bars show the fold change in abundance of [1,1,2,3,0]. The white bars show the fold change in abundance of [0,2,2,3,0]. Error bars are calculated from triplicate experiments and are reported as +/− S.D.

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.
3.
Figure 9

Figure 9. Comparison of the extent of HSulf2 mediated release of 6O-sulfate from bovine and murine HS samples.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

The release of 6O-sulfate from the various HS samples was determined by summing the decreases in abundance of 6O-sulfated disaccharides (Fig. 1A and Fig. 5A). (A) The release of 6O-sulfate by HSulf2 as a function of total 6O-sulfate content in the control samples. (B) The release of 6O-sulfate from D2S6 after HSulf2 treatment as a function of total D2S6 content in the control samples.

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.
4.
Figure 6

Figure 6. Fold change plot of the abundances of saturated disaccharides from control and HSulf2 treated murine HS samples.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

Control or HSulf2 treated murine HS samples were analyzed using SEC-MS. Changes in disaccharide abundance after HSulf2 treatment are shown as a fold-change difference. The changes in the abundance of U0A6 and U0S6 with respect to their isobars cannot be determined due to the low abundance of saturated disaccharides. Error bars are calculated from triplicate experiments and are reported as +/− S.D.

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.
5.
Figure 3

Figure 3. Fold change plot of the abundances of saturated disaccharides from control and HSulf2 treated bovine HS samples.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

Control or HSulf2 treated bovine HS samples were analyzed using SEC-MS. Changes in disaccharide abundance after HSulf2 treatment are shown as a fold-change difference. The changes in the abundance of U0A6 and U0S6 with respect to their isobars cannot be determined due to the low abundance of saturated disaccharides. Error bars are calculated from triplicate experiments and are reported as +/− S.D.

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.
6.
Figure 7

Figure 7. Influence of HSulf2 digestion on Δ4,5-unsaturated and saturated disaccharide abundances following lyase depolymerization.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

The abundances of total saturated and Δ4,5-unsaturated disaccharides in control or HSulf2 treated samples were compared and expressed as a LN fold change. (A) The fold-change in abundance of Δ4,5-unsaturated (black bars) or saturated (white bars) disaccharides subsequent to HSulf2 treatment of bovine HS samples. (B) The fold-change in abundance of Δ4,5-unsaturated (black bars) or saturated (white bars) disaccharides subsequent to HSulf2 treatment of murine HS samples. Error bars are calculated from triplicate experiments and are reported as +/− S.D.

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.
7.
Figure 4

Figure 4. Changes in degree of sulfation of bovine HS NS domains due to HSulf2 digestion.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

The maximum possible change in the degree of sulfation (sulfates per disaccharide unit) of NS domains from bovine HS chains after HSulf2 treatment (black bars) was calculated, based on previously quantified nitrous acid depolymerization products generated from the same sample pool [11]. This is compared to the change in degree of sulfation calculated based on the disaccharide data generated during the current study (Fig. 1A).

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.
8.
Figure 5

Figure 5. Fold change plot of the abundances of Δ4,5-unsaturated disaccharides from control and HSulf2 treated murine HS samples. Control or HSulf2 treated murine HS samples were analyzed using SEC-MS.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

(A) Disaccharide abundances after HSulf2 treatment are shown as a fold-change difference relative to control samples. Two other 6O-sulfated disaccharides, D0A6 and D0S6, occur as isomer pairs with D2A0 and D2S0, respectively. Tandem MS was used to determine the percent abundance of each 6O-sulfated disaccharide (within its isobar), before and after HSulf2 treatment. (B) The relative abundance of D0A6 before and after HSulf2 treatment. (C) The relative abundance of D0S6 before and after HSulf2 treatment. Error bars are calculated from triplicate experiments and are reported as +/− S.D.

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.
9.
Figure 1

Figure 1. Disaccharide abundances before and after HSulf2 digestion.. From: Glycomics Analysis of Mammalian Heparan Sulfates Modified by the Human Extracellular Sulfatase HSulf2.

(A) Abundances of Δ4,5-unsaturated disaccharide abundances from control and HSulf2 treated HS. Peak areas were determined by integration of extracted ion chromatograms for each of the structures indicated on the X axis. Error bars are calculated from triplicate experiments and are reported as +/− S.D. Two 6O-sulfated disaccharides, D0S6 and D0A6, occur as isomer pairs with D2S0 and D2A0, respectively. Tandem MS was used to determine the percent abundance of each 6O-sulfated disaccharide (within its isobar), before and after HSulf2 treatment. (B) The percent abundance of D0S6 in the D2S0/D0S6 isomer pair before and after HSulf2 treatment. (C) The relative abundance of D0A6 in the D2A0/D0A6 isomer pair before and after HSulf2 treatment.

Gregory O. Staples, et al. PLoS One. 2011;6(2):e16689.

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