TABLE 1.
Concentrations of MgATP and ATP required for half-maximal binding by wild-type and mutant forms of NtrC
| Protein | ATPase activitya (%) | Concn for half-maximal binding (μM)b
|
|
|---|---|---|---|
| MgATP | ATP | ||
| Wild type | 100c | 135d ± 10 (6) | 730 ± 120 (3) |
| NtrCG173N | ≤2 | ≥10,000 (3) | ND |
| NtrCS207F | ≤2 | 50 ± 5 (4) | 230 ± 25 (3) |
| NtrCE208Q | 12 | 105 ± 10 (4) | 400 (2) |
| NtrCA216V | 100 | 100 ± 10 (3) | 350 (1) |
| NtrCG219K | 56 | 40 ± 10 (3) | 510 (2) |
| NtrCA220T | 80 | 100 ± 10 (4) | 300 (1) |
| NtrCD239N | ≤2 | 20 ± 5 (3) | 40 ± 15 (3) |
| NtrCD239C | ≤2 | 25 ± 5 (3) | 20 ± 5 (3) |
| NtrCD239A | ≤2 | 30 ± 10 (3) | 10 ± 5 (3) |
| NtrCR294C | ≤2 | 190 ± 35 (3) | 850 (1) |
| NtrCG355V | ≤2 | 800 ± 250 (3) | ND |
| NtrCR358C | ≤2 | ≥10,000 (3) | ND |
| NtrCR358H | ≤2 | ≥10,000 (2) | ND |
From references 67 and 42, with the exception of data for proteins carrying substitutions at position 239. Proteins were phosphorylated.
Binding by unphosphorylated proteins was assessed as described in Materials and Methods, and titration data were plotted to determine concentrations for half-maximal binding (see Materials and Methods). The number of independent titrations carried out for each NtrC protein (usually 10 μM monomer; range from 7 to 20 μM monomer in occasional experiments) is shown in parentheses, and standard deviations are indicated where appropriate. ND, not determined.
100% corresponds to ∼500 pmol/15 min/10 μl at a protein concentration of 1 μM.
Weighted average for two independently purified wild-type NtrC preparations. For the remaining proteins, single preparations were titrated.