Table 1 Analysis of expression and ATPase activity with PS and PE.

From: Disease mutations reveal residues critical to the interaction of P4-ATPases with lipid substrates

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

  1. 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).
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