FIGURE 8.

RPIP-HPLC chromatograms of the disaccharide analysis of the polysaccharides modified by various C₅-epi mutants and 2OST WT. The polysaccharides were prepared by incubating with C₅-epi and C₅-epi point mutants followed by the [³⁵S]sulfation using 2OST WT. The resultant disaccharides were resolved using RPIP-HPLC, as described under “Experimental Procedures.” A–D, chromatograms of the polysaccharide treated with C₅-epi WT (A), C₅-epi Y146A (B), C₅-epi Y168A (C), and no C₅-epi (D), respectively. It should be noted here that in C and D, the presence of the small peak at ∼49 min may be an artifact from nitrous acid degradation, and no IdoUA has been found in N-sulfoheparosan.