Table 3. Analysis of NrfA and NrfB interactions by AUC.
Sedimentation experiments were performed on NrfA, and mixtures of NrfA and NrfB over a range of concentrations using a Beckman XL-I analytical centrifuge. NrfA was centrifuged at 9000 rev./min, while the NrfA–NrfB complex was centrifuged at 7500 and 9000 rev./min. Scans were collected every 4 h until equilibrium was achieved. The averaged molecular mass was determined for each sample using Ultrascan II [23]. The Kd was determined for NrfA and the NrfA–NrfB complex assuming monomeric molecular masses of 53 and 74 kDa respectively. The molecular mass shown for NrfA–NrfB samples is the averaged molecular mass determined at the two different speeds.
| Sample (μM) | λ (nm) | ϵ (mM−1·cm−1) | Averaged molecular mass (kDa) | Kd (μM) |
|---|---|---|---|---|
| NrfA (2.0) | 410 | 497 | 72 | |
| NrfA (10) | 440 | 110 | 83 | 4.0 |
| NrfA (20) | 530 | 48 | 98 | |
| NrfA–NrfB (1.0) | 410 | 1000 | 97±4 | |
| NrfA–NrfB (5.0) | 530 | 96 | 134±1 | 4.2 |
| NrfA–NrfB (20) | 600 | 15 | 147±9 |