Table 1.
Kinetic Constants for ATP Hydrolysis for PspF1-275 WT and Variants
| Protein | Vmax (min−1) | Km (μM)a |
|---|---|---|
| WT | 39.51 ± 4.62 | 250 |
| L9A | 20.72 ± 2.55 | 2,300 |
| K42A | Not detected | Not detected |
| E43A | 1.37 ± 0.10 | 750 |
| E43D | 22.26 ± 4.80 | 250 |
| E43Y | 2.49 ± 0.17 | 1,250 |
| L44A | 18.16 ± 2.42 | 310 |
| N64Q | 1.50 ± 0.10 | 500 |
| D107A | 0.23 ± 0.05 | 300 |
| E108A | 0.15 ± 0.01 | 100 |
| E108Q | 0.07 ± 0.01 | 100 |
| E118A | 0.60 ± 0.05 | >1,500 |
| E118D | 20.23 ± 3.70 | 800 |
| E118R | 0.31 ± 0.05 | 50 |
| R122A | 1.26 ± 0.03 | 300 |
| R122E | 0.95 ± 0.06 | 200 |
| E125A | 8.86 ± 0.38b | 2,000 |
| E125D | 0.43 ± 0.01 | 160 |
| E125Q | 3.39 ± 0.38b | 170 |
| Y126A | 11.95 ± 1.77 | 650 |
| Y126E | 12.82 ± 1.03 | 800 |
| R162A | 0.18 ± 0.02 | 90 |
| R162E | 0.28 ± 0.01 | >3,000 |
| R162K | 1.88 ± 0.29 | 200 |
| R162H | 0.08 ± 0.01 | 3,000 |
| D164A | 0.21 ± 0.02 | 120 |
| D164N | 1.83 ± 0.13c | 155 |
| D164Q | 0.13 ± 0.04 | >5,000 |
| R168A | 0.17 ± 0.03 | >4,500 |
| R168E | 0.11 ± 0.03 | >3,000 |
| R168K | 0.12 ± 0.02 | >4,000 |
| R168H | 0.43 ± 0.06 | >5,000 |
a
Michaelis-Menten kinetic constants for WT and mutated PspF1-275 variants in the absence of σ54. The data were an average of at least three independent experiments. Maximal error is 10%.
b
No Vmax, linear with concentration/no plateau.
c
Max already at low concentration/no sigmoidal curve.