TABLE 3.
The effect of mutation of charged residues on Phormidium PC in the presence of the Zn2+ ion on its interaction with Phormidium cyt f
| Ionic strength (mM)
|
|||||||
|---|---|---|---|---|---|---|---|
| 10
|
100
|
||||||
| Class* | Mutant† | Complexes formed per 1000 trajectories† | % WT | 10−8 × k2 (M−1 s−1)† | Complexes formed per 1000 trajectories† | % WT | 10−8 × k2 (M−1 s−1)† |
| WT | 227 ± 11 | 100 | 56 ± 3 | 39.2 ± 0.8 | 100 ± 3 | 11.6 ± 0.8 | |
| II | 130–200% WT complex formation at 10 mM ionic strength | ||||||
| D45(43)A‡ | 361 ± 22 | 160§ | 83 ± 3 | 76.3 ± 1.2 | 191 ± 5 | 21.3 ± 0.7 | |
| D57(55)A | 335 ± 16 | 149 | 78 ± 3 | 56.8 ± 1.3 | 143 ± 5 | 16.1 ± 1.0 | |
| D44(42)A | 321 ± 7 | 143 | 78 ± 3 | 71.8 ± 0.9 | 180 ± 4 | 20.0 ± 1.0 | |
| E54(52)A | 317 ± 15 | 141 | 78 ± 3 | 50.0 ± 2.8 | 127 ± 8 | 14.0 ± 0.9 | |
| D10(8)A | 314 ± 8 | 140 | 74 ± 3 | 47.3 ± 3.6 | 120 ± 9 | 13.5 ± 1.0 | |
| E17(15)A | 310 ± 7 | 138 | 73 ± 3 | 46.0 ± 0.3 | 117 ± 3 | 13.1 ± 0.7 | |
| E70(68)A | 303 ± 5 | 135 | 72 ± 3 | – | – | – | |
| III | 100–133% WT complex formation at 10 mM ionic strength | ||||||
| E73(71)A | 278 ± 4 | 124 | 67 ± 3 | – | – | – | |
| D27(25)A | 252 ± 8 | 111 | 61 ± 3 | – | – | – | |
| E104(99)A | 246 ± 12 | 108 | 60 ± 4 | – | – | – | |
| E1(−3)A | 244 + 11 | 108 | 61 ± 4 | – | – | – | |
| D79(77)A | 241 ± 12 | 107 | 59 ± 3 | 41.8 ± 1.3 | 107 ± 4 | 11.8 ± 0.9 | |
| IV | 40–100% WT complex formation at 10 mM ionic strength | ||||||
| K30(28)A | 169 ± 8 | 75 ± | 44 ± 3 | 37.2 ± 1.3 | 95 ± 4 | 10.7 ± 0.7 | |
| K58(56)A | 162 ± 6 | 72 ± | 42 ± 3 | 34.3 ± 1.3 | 88 ± 4 | 9.8 ± 0.9 | |
| K6(4)A | 151 ± 12 | 67 | 39 ± 3 | 35.0 ± 1.1 | 90 ± 3 | 10.0 ± 0.8 | |
| K35(33)A | 141 ± 10 | 63 | 37 ± 3 | 31.8 ± 0.8 | 82 ± 3 | 9.2 ± 0.9 | |
| K100(95)A | 121 ± 5 | 54 | 32 ± 3 | – | – | – | |
| K53(51)A | 114 ± 7 | 51 | 30 ± 2 | 25.3 ± 0.7 | 66 ± 2 | 7.2 ± 0.6 | |
| V | <40% WT at 10 mM ionic strength | ||||||
| K46(44)A | 71 ± 11 | 32 | 20 ± 2 | 20.7 ± 1.0 | 54 ± 3 | 6.0 ± 0.6 | |
| R93(88)A | 55 ± 7 | 25 | 15 ± 3 | 13.4 ± 0.4 | 37 ± 1 | 3.9 ± 0.5 | |
| R93(88)Q | 51 ± 10 | 23 | 14 ± 2 | 14.9 ± 0.5 | 40 ± 2 | 4.3 ± 0.5 | |
The number of complexes formed in the absence of the electrostatic field (2.2/1000 trajectories) was subtracted from the total number of complexes formed to determine the electrostatic contribution. The mutants were divided into classes based on their effects on complex formation. Class I, significant stimulation of complex formation (i.e., the number of complexes formed was >200% WT at 10 mM ionic strength; class II, 130–200% WT at 10 mM; class III, the number of complexes formed was between 100 and 133% WT at 10 mM; class IV, the number of complexes formed was between 40 and 100% WT at 10 mM; and class V, the number of complexes formed was <40% WT at 10 mM ionic strength. Five sets each of 1000 and 5000 trajectories were carried out at 10 and 100 mM, respectively.
The mutants were made as described in the Methods section. Rates were determined as described in the Methods section and for Table 1. The rate obtained in the absence of the electrostatic field (0.6 × 108 M−1 s−1) was subtracted from the total rate to determine the electrostatic contribution.
The numbers in the parentheses refer to the consensus sequence of Pearson (2000).
Standard deviations averaged 5% of the values shown.