Table 1.
DHPAAS and DDC parameters of enzyme variants included in this study
| B. mori DHPAAS variant | DHPAAS Km: µM L-DOPA | DHPAAS kcat: min−1 | DHPAAS Vmax: nmol H2O2 min−1 mg−1 | DDC Vmax: nmol dopamine min−1 mg−1 | Estimated % DHPAAS activity |
|---|---|---|---|---|---|
| WT DHPAAS | 41.7 (36.6–47.3) | 6.51 (6.27–6.76) | 115 | 5.59 | 95% |
| Phe79Tyr-Tyr80Phe | 62.3 (49.5–78.5) | 4.65 (4.30–5.03) | 82.2 | 41.4 | 67% |
| Asn192His | 36.5 (32.8–40.5) | 1.40 (1.35–1.44) | 24.7 | 87.6 | 22% |
| Phe79Tyr-Tyr80Phe- Asn192His | 27.6 (17.1–42.8) | 0.97 (0.87–1.09) | 15.4 | 247 | 5.9% |
Km, kcat, and Vmax values were calculated with Prism 7 using H2O2 production data from Fig. 6. 95% confidence intervals listed in parenthesis (n ≥ 8 independent measurements). DDC activity Vmax is calculated based on the first in vitro measurements of dopamine product (Fig. 7a, t = 9 min.). Estimated % DHPAAS activity is based on the comparison of these two rates.