TABLE 3.
Kinetic constants of 2,3-dihydroxybiphenyl 1,2-dioxygenases BphC1, BphC2, and BphC3 with monochloro-2,3-dihydroxybiphenyls
| Substrate and enzyme | kcat (s−1) | Km (μM) | Kssa (μM) | kcat/Km (s−1 mM−1) | Partition ratio |
|---|---|---|---|---|---|
| 2,3-Dihydroxybiphenyl | |||||
| BphC1bb | 4.8 ± 0.1d | 1.0 ± 0.1 | 3,300 ± 900 | 4,800 | 1,410 ± 240 |
| BphC2c | 16.5 ± 2.1 | 25.6 ± 5.1 | 340 ± 50 | 640 | 2,250 ± 200 |
| BphC3 | 9.8 ± 0.3 | 29.4 ± 3.1 | 2,860 ± 550 | 330 | 1,930 ± 110 |
| 2′-Chloro-2,3-dihydroxybiphenyl | |||||
| BphC1b | 0.25 ± 0.01 | 1.2 ± 0.1 | 5,550 ± 650 | 170 | 470 ± 70 |
| BphC2c | 7.5 ± 0.4 | 18.7 ± 1.0 | >105 | 400 | 650 ± 80 |
| BphC3 | 3.5 ± 0.4 | 18.3 ± 1.5 | >105 | 190 | 850 ± 90 |
| 3′-Chloro-2,3-dihydroxybiphenyl | |||||
| BphC1b | 4.0 ± 0.1 | 0.45 ± 0.05 | 5,300 ± 360 | 8,900 | 460 ± 90 |
| BphC2c | 12.0 ± 1.0 | 9.7 ± 1.1 | >105 | 1,200 | 1,120 ± 60 |
| BphC3 | 8.5 ± 2.0 | 9.9 ± 0.9 | >105 | 860 | 1,190 ± 110 |
| 4′-Chloro-2,3-dihydroxybiphenyl | |||||
| BphC1b | 4.5 ± 0.2 | 1.5 ± 0.1 | 6,100 ± 1,300 | 3,000 | 270 ± 20 |
| BphC2c | 18.8 ± 1.5 | 10.6 ± 1.1 | >105 | 1,800 | 2,490 ± 420 |
| BphC3 | 8.3 ± 0.3 | 12.0 ± 0.7 | >105 | 690 | 1,050 ± 80 |
a
Kss, substrate inhibition constant (20).
b
Values are given relative to protein content. As BphC1 contains only about 12% ± 4% Fe2+ per subunit, the values of kcat, kcat/Km, and the partition ratio can be assumed to be underestimated by a factor of 6 to 12.
c
Values are given relative to protein content. As BphC2 contains about 70% ± 5% Fe2+ per subunit, the values of kcat, kcat/Km, and the partition ratio can be assumed to be underestimated by a factor of 1.4.
d
Mean±standard deviation.