FIGURE 7.

Anion-exchange HPLC chromatograms of oligosaccharide substrates modified by 2OST wild type and mutant proteins. Two oligosaccharide substrates, compound 3 and 4, were incubated with 2OST proteins in the presence of ³⁵S-labeled PAPS followed by anion-exchange HPLC analysis. A–C represent chromatograms of the GlcA-containing substrate incubated with wild type 2OST (red tracing), 2OST R189A, and 2OST R354A, respectively. D–F represent chromatograms of the IdoA-containing substrate incubated with wild type 2OST, 2OST R189A, and 2OST R354A, respectively. Control reactions have no enzyme (blue tracing in both A and D). For the left column (A–C), the ³⁵S-labeled peak eluted at 32–33 min contains the desired product. The ³⁵S-labeled peak eluted at 38 min is PAPS. * indicates an unidentified peak. For the right column (D–F), the³⁵S-labeled peak eluted at 35–36 min is the desired product.