2.5 Hydrolysis enzymatic synthesis and fermentation
Enzyme reactions were performed based on the conditions in Table 3.
Furthermore, reaction conditions were investigated, including
temperature, pH value, buffer type, concentration, and the Ps XR
and Pd PDHD36A/I37R ratio, as shown in Figure
S1. WP was hydrolyzed in citrate buffer at pH 5.5, 55 °C with a suitable
amount of β -galactosidase. Enzymatic hydrolysis was performed in
a conical flask containing 122 g/L WP or 100 g/L lactose, 50 mM citrate
buffer (pH 5.5, 55 °C), and a suitable amount of β -galactosidase
for 3 h. Enzymatic synthesis and fermentation were performed in a
conical flask containing previously obtained enzymatic hydrolysates, 100
mM Tris-HCl buffer (pH 8.0, 30 °C), suitable amounts ofPd PDH-Ps XR complex, cofactor NADPH, and yeast cells with a
final A600 = 1. For the fermentation of enzymatic
hydrolysates derived from lactose, 2% tryptone was required. The
protein content of the WP solution and yeast cells was determined using
a BCA Protein Assay Kit, as previously
reported.[14]