Table 1. Kinetic Parameters for Nitrite Reduction Catalyzed by YtfE.
| mediator | potential (mV) versus SHE | pH | YtfEa | Km (μM NO2–) | Vmaxb (μmol min–1 mg–1) | kcat (min–1) | kcat/Km (M–1 min–1) |
|---|---|---|---|---|---|---|---|
| Fe2+(RS)n | –330 | 7.5 | wt | 88 (± 13) | 0.04 (± 0.001) | 0.97 (± 0.04) | 1.10 (± 0.044) × 104 |
| 7.5 | wt + O2 | 242 (± 81) | 0.02 (± 0.003) | 0.56 (± 0.07) | 0.23 (± 0.028) × 104 | ||
| 7.5 | C30A/C31A | n.d. | ∼0.001 | ∼0.03 | n.d. | ||
| 7.5 | CAM-wt | n.d. | ∼0.002 | ∼0.05 | n.d. | ||
| safranin O | –289 | 7.5 | wt | 252 (± 32) | 0.22 (± 0.01) | 5.74 (± 0.20) | 2.28 (± 0.080) × 104 |
| 7.5 | C30A/C31A | 251 (±32) | 0.23 (± 0.01) | 6.11 (± 0.30) | 2.43 (± 0.119) × 104 | ||
| 7.5 | CAM-wt | 252 (± 32) | 0.23 (± 0.01) | 6.12 (± 0.38) | 2.43 (± 0.150) × 104 | ||
| methyl viologen | –450 | 7.0 | wt | 279 (± 70) | 2.40 (± 0.23) | 62.7 (± 5.95) | 22.49 (± 2.132) × 104 |
| 7.5 | wt | 252 (± 32) | 1.33 (± 0.01) | 34.8 (± 1.59) | 13.81 (± 0.631) × 104 | ||
| 7.5 | C30A/C31A | 288 (± 29) | 1.59 (± 0.01) | 41.6 (± 1.58) | 14.44 (± 0.550) × 104 | ||
| 8.0 | wt | 246 (± 32) | 0.76 (± 0.03) | 19.8 (± 0.82) | 8.04 (± 0.333) × 104 |
a
Note that wt is wild-type YtfE and CAM-wt is carboxymethylated YtfE.
b
Mediator-dependent data, expressed as NO-generated (μmol min–1 mg–1) (YtfE), safranin O-oxidized (μmol min–1 mg–1), or methyl viologen-oxidized (μmol min–1 mg–1), respectively.