Table 1.
Analysis of expression and ATPase activity with PS and PE.
| Mutanta | Expressionb | ATPase activity stimulated by PSc | ATPase activity stimulated by PEc | |||||||
|---|---|---|---|---|---|---|---|---|---|---|
| % of WT | n | Vmax with PS, µmol/min/mg | n | K0.5 for PS, µM | n | Vmax with PE, µmol/min/mg | n | K0.5 for PE, µM | n | |
| WT | 100 ± 12 | 14 | 97 ± 4 | 31 | 34 ± 1 | 342 | 50 ± 3 | 26 | 386 ± 16 | 264 |
| I91P (L127P) | 12 ± 2*** | 4 | 18 ± 2 (5.4x↓)*** | 6 | 96 ± 6 (2.8x↑)*** | 60 | 3 ± 0.4 (17x↓)*** | 2 | 1245 ± 522 (3.2x↑)*** | 20 |
| I91A | 29 ± 3** | 3 | 88 ± 15 (1.1x↓) | 7 | 65 ± 4 (1.9x↑)*** | 70 | 46 ± 9 (1.1x↓) | 4 | 516 ± 48 (1.3x↑) | 40 |
| L308F (I344F) | 104 ± 34 | 2 | 85 ± 1 (1.1x↓) | 4 | 151 ± 10 (4.4x↑)*** | 40 | 28 ± 2 (1.8x↓)** | 4 | 923 ± 65 (2.4x↑)*** | 40 |
| L308A | 88 ± 18 | 2 | 96 ± 5 (1.0x) | 5 | 38 ± 2 (1.1x↑) | 50 | 35 ± 4 (1.4x↓) | 4 | 430 ± 18 (1.1x↑) | 39 |
| E897K (E981K) | 29 ± 7* | 2 | 19 ± 4 (5.1x↓)*** | 8 | 11 ± 1 (3.1x↓)*** | 78 | 9 ± 3 (5.6x↓)*** | 5 | 422 ± 48 (1.1x↑) | 50 |
| E897A | 54 ± 18 | 3 | 41 ± 8 (2.4x↓)*** | 8 | 17 ± 1 (2.0x↓)*** | 79 | 33 ± 8 (1.5x↓)** | 10 | 349 ± 30 (1.1x↓) | 104 |
| E897D | 98 ± 5 | 2 | 92 ± 3 (1.1x↓) | 4 | 43 ± 3 (1.3x↑) | 40 | 50 ± 7 (1.0x) | 4 | 495 ± 31 (1.3x↑) | 39 |
| E897Q | 78 ± 10 | 3 | 60 ± 3 (1.6x↓)* | 5 | 17 ± 1 (2.0x↓)*** | 50 | 38 ± 13 (1.3x↓) | 5 | 255 ± 15 (1.5x↓) | 50 |
| E897F | 45 ± 7 | 2 | 31 ± 4 (3.1x↓)*** | 4 | 7 ± 1 (4.9x↓)*** | 39 | 71 ± 12 (1.4x↑)* | 4 | 476 ± 28 (1.2x↑) | 39 |
| E897R | 29 ± 1* | 2 | 10 ± 1 (9.7x↓)*** | 4 | 13 ± 1 (2.6x↓)*** | 40 | 3 ± 0.1 (17x↓)*** | 5 | 1205 ± 59 (3.1x↑)*** | 50 |
| R898A | 66 ± 8 | 5 | 76 ± 12 (1.3x↓) | 8 | 53 ± 3 (1.6x↑)*** | 80 | 27 ± 3 (1.9x↓)*** | 9 | 353 ± 30 (1.1x↓) | 90 |
| R898D | 89 ± 12 | 3 | 83 ± 11 (1.2x↓) | 5 | 126 ± 4 (3.7x↑)*** | 60 | 9 ± 0.3 (5.6x↓)*** | 4 | 1069 ± 140 (2.8x↑)*** | 39 |
| R898E | 82 ± 13 | 2 | 59 ± 13 (1.6x↓)** | 6 | 97 ± 5 (2.9x↑)*** | 70 | 12 ± 4 (4.2x↓)*** | 5 | 719 ± 93 (1.9x↑)*** | 50 |
| R898K | 79 ± 5 | 2 | 105 ± 22 (1.1x↑) | 4 | 30 ± 1 (1.1x↓) | 50 | 43 ± 7 (1.2x↓) | 4 | 591 ± 49 (1.5x↑) | 50 |
| E897R_R898E | 56 ± 1 | 2 | 35 ± 2 (2.8x↓)*** | 4 | 52 ± 3 (1.5x↑)*** | 40 | 11 ± 5 (4.5x↓)*** | 4 | 888 ± 37 (2.3x↑)*** | 40 |
| D99A | 85 ± 2 | 2 | 85 ± 14 (1.1x↓) | 6 | 40 ± 1 (1.2x↑) | 40 | 17 ± 4 (2.9x↓)*** | 4 | 541 ± 29 (1.4x↑) | 40 |
| R105A | 62 ± 9 | 2 | 48 ± 6 (2.0x↓)** | 4 | 45 ± 3 (1.3x↑)** | 79 | 25 ± 2 (2.0x↓)* | 3 | 487 ± 47 (1.3x↑) | 27 |
Data deviating significantly from WT according to the one-way ANOVA test (see further in Methods) are marked with *(0.05 > P > 0.01), **(0.001 < P < 0.01), and ***(P < 0.001). aWild type (WT) and mutant bATP8A2 proteins. Homologous disease mutations in human ATP8B1 are indicated in parentheses. bAverage of expression levels calculated relative to the wild type of the same transfection experiment, n indicating the number of transfection experiments. cVmax refers to the ATPase activity at the highest lipid concentration examined (1000 µM PS or 2000 µM PE), K0.5 (substrate concentration giving half maximum activation) was obtained by nonlinear regression curve fitting (see Figs 3 and 4), n indicating the number of data points in the regression. In parentheses is shown fold change relative to wild type (arrow pointing upward/downward for increase/decrease).