*2.3. Kinetic Study of the Biotransformation of GAA by BsGT110*

To study how pH affects the biotransformation activity of GAA by BsGT110, a kinetic study of the biotransformation was conducted at different concentrations of GAA, with 50 mM acetate buffer at pH 6 or PB at pH 7, 10 mM MgCl2, and 10 mM UDP-glucose, at 40 ◦C. The results of the kinetic study were shown in Figure 5, and the calculated kinetic parameters were listed in Table 2. The results showed that the turnover number (kcat) of BsGT110 toward GAA at pH 6 was 3-fold higher than that at pH 7.

**Figure 5.** Kinetic study of BsGT110 at pH 6 (closed symbols) and pH 7 (open symbols). Different concentrations of GAA were mixed with 15 μg purified BsGT110 protein, 10 mM UDP-glucose, 10 mM MgCl2, and 50 mM PB (pH 7.0) or acetate buffer (pH 6) in 1 mL reaction mixture and incubated at 40 ◦C for 20 min. During the incubation, samples from each reaction were removed and analyzed by UPLC every 2 min. The reaction rate for each concentration of GAA was obtained from the slope of the plot of the amount of product over time. The UPLC operation procedure was described in the Materials and Methods section.


**Table 2.** Kinetic parameters of BsGT110 toward GAA at pH 6 and pH 7.
