FIGURE 4.

Dose response of C₅-epi to the [³⁵S]sulfation by 2OST Y94I and 2OST WT. Different amounts of C₅-epi (0–66 μg/ml) were incubated with 2.0 μg of N-sulfoheparosan for 30 min at 37 °C. Then epimerization reactions were stopped by boiling for 5 min. After centrifugation, the supernatants were mixed with either 2OST WT or 2OST Y94I and [³⁵S]PAPS (6.4 × 10⁵ cpm), followed by incubation for 5 min at 37 °C. The sulfated polysaccharide was purified by DEAE resin and determined by scintillation counting. The results (average ± S.D.) obtained using 2OST WT are shown by filled circles, and 2OST Y94I results are shown by empty circles. The bars indicate S.E. of triplicates.