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Table. 2.

Table. 2.

Michaelis-Menten constant (Km), maximum velocity (Vmax), the catalytic rate constant (kcat), and the coupling efficiency using purified CYP154C3s, PdR, Pdx, and 10 different steroid substrates.

SN Substrate Km [μM] Vmax [μM Product / μM CYP / min] Kcat [1/S] Coupling efficiency (%)

CYP154C3-1 CYP154C3-2 CYP154C3-1 CYP154C3-2 CYP154C3-1 CYP154C3-2 CYP154C3-1 CYP154C3-2
1 Adrenosterone 30.110 ± 1.711 27.850 ± 1.820 0.804 ± 0.013 0.793 ± 0.017 48.240 ± 0.780 47.580 ± 1.020 29.440 ± 3.770 31.800 ± 4.380
2 Androstenedione 38.480 ± 4.350 26.980 ± 1.540 0.844 ± 0.031 0.787 ± 0.014 50.640 ± 1.860 47.220 ± 0.840 27.430 ± 3.920 22.040 ± 3.830
3 Corticosterone 59.700 ± 6.350 81.170 ± 11.630 0.941 ± 0.031 0.923 ± 0.046 56.460 ± 2.100 55.380 ± 2.760 19.380 ± 3.280 19.880 ± 3.300
4 Cortisone 83.500 ± 6.670 108.510 ± 12.480 0.930 ± 0.027 0.979 ± 0.041 55.800 ± 1.620 58.740 ± 2.460 21.650 ± 3.390 23.410 ± 3.920
5 Hydrocortisone 141.420 ± 18.120 170.750 ± 22.780 0.231 ± 0.015 0.228 ± 0.015 13.860 ± 1.600 13.680 ± 1.900 13.320 ± 2.860 11.870 ± 3.060
6 Nandrolone 53.820 ± 7.170 49.830 ± 11.180 0.893 ± 0.043 0.821 ± 0.068 53.580 ± 2.280 49.260 ± 4.050 23.530 ± 3.470 24.990 ± 3.590
7 Prednisolone 246.540 ± 38.310 280.990 ± 37.510 0.294 ± 0.023 0.285 ± 0.020 17.640 ± 1.380 17.100 ± 1.200 8.770 ± 3.140 9.160 ± 2.720
8 Prednisone NA NA NA NA NA NA 25.130 ± 2.460 24.230 ± 3.750
9 Progesterone 6.320 ± 0.710 10.670 ± 1.220 0.833 ± 0.210 0.765 ± 0.029 49.980 ± 1.260 45.900 ± 1.740 39.490 ± 4.120 36.060 ± 4.280
10 Testosterone 10.380 ± 0.830 18.260 ± 3.220 0.933 ± 0.025 0.915 ± 0.052 55.980 ± 1.500 54.900 ± 3.120 35.440 ± 4.320 33.680 ± 4.170

The overall apparent kinetic parameters were determined with a CYP: Pdx: Pdr concentration ratio of 1: 8: 2 for the purified CYP154C3s toward 10 substrates. Coupling efficiency was calculated as the percentage of NADH used for the formation of product over the total NADH consumption. The NADH consumption rate was calculated after subtracting the respective background NADH consumption. The results represent the mean values of triplicate measurements.

J. Microbiol. Biotechnol. 2021;31:464~474 https://doi.org/10.4014/jmb.2010.10020
© J. Microbiol. Biotechnol.