Fig. 2. Increasing enzyme concentration relative to substrate alters the selectivity of 3-OST-3, enabling sulfation of a 6-O-sulfated substrate (8-mer 3). (A) Titration of 3-OST-3 enzyme concentration using either 8-mer 1 (black bar) or 8-mer 3 (grey bar) at constant substrate concentration. Both reactions show increased progression, measured as pmoles of sulfo groups transferred, with increasing enzyme to substrate ratio. This suggests both oligosaccharides are viable substrates for 3-OST-3. (B) Graph depicting the trend in sulfo groups transferred with increasing enzyme concentration for both 8-mer 1 (open circle) and 8-mer 3 (closed square). As enzyme concentration increases, the difference between the 8-mer 1 and 8-mer 3 lines decreases, suggesting the selectivity of 3-OST-3 toward non-6-O-sulfated substrates is enzyme-concentration dependent.