2.3. Whole-cell reaction
The harvested BL21 Star (DE3) cells expressing LCD were used for the
whole-cell reaction. For l-PA production, 200 mM
l-lysine was used as substrate, and 150 mM potassium phosphate
buffer (pH 7) was used to buffer the mixture. To enhance cell
permeability, 0.1% triton X-100 was added. FeSO4 at 20
mM was added as source of iron. Distilled water was added to the
reaction mixture containing harvested cells with an
OD600 of 50 to prepare a total volume of 400 μL. The
mixture was titrated to pH 7.5 with 2 N NaOH. The reaction was conducted
at 37 °C with gentle shaking (40–50 rpm). After reaction, 100 μL of
each sample was collected and heated at 95 °C for 5 min to complete the
reaction. Inactivated cells were eliminated by centrifugation at 13,000
rpm, and the supernatant was diluted to an appropriate concentration for
analysis.