Construct | Residue2-a | EC50 | ||
---|---|---|---|---|
2MeSATP | 2MeSADP | HT-AMP | ||
nM | ||||
Wild-type | 6.75 ± 2.58 | 1.27 ± 0.01 | 355 ± 4 | |
R128A | 3.29 | 2-b | 2-b | 2-b |
F131A | 3.32 | 44.2 ± 15.9 | 18.2 ± 2.1 | 5260 ± 2200 |
H132A | 3.33 | 79.9 ± 29.6 | 15.9 ± 4.4 | 2390 ± 600 |
Y136A | 3.37 | 119 ± 103 | 12.6 ± 1.8 | 1720 ± 690 |
T221A | 5.42 | 604 ± 456 | 15.9 ± 4.0 | 1810 ± 280 |
T222A | 5.43 | 221 ± 58 | 11.2 ± 0.6 | 1210 ± 210 |
F226A | 5.47 | 56.8 ± 7.5 | 20.7 ± 3.8 | 1850 ± 800 |
H277A | 6.52 | 115 ± 28 | 100 ± 30 | 4520 ± 640 |
K280A | 6.55 | 6400 ± 1380 | 1790 ± 310 | 203,000 ± 36,000 |
Q307A | 7.36 | 1400 ± 300 | 455 ± 140 | 43,000 ± 600 |
R310A | 7.39 | 2-b | 2-b | 2-b |
R310K | 7.39 | 4580 ± 430 | 418 ± 118 | 13,400 ± 1700 |
R310S/S314R | 7.39/7.43 | 2-b | 2-b | 2-b |
S314A | 7.43 | 2-b | 2-b | 2-b |
S314T | 7.43 | 60.0 ± 18.0 | 11.7 ± 1.8 | 1950 ± 440 |
S317A | 7.46 | 8.00 ± 1.78 | 1.58 ± 0.29 | 611 ± 52 |
Data are presented as means ± standard deviation of two to four independent experiments, each performed in duplicate. All constructs contain the Ha-epitope tag sequence at the amino terminus (5) and a hexa-His tag at the carboxyl terminus (25).