TABLE 3.
Apparent affinities for cytoplasmic and luminal Ca2+
K0.5 ± S.E. values were obtained by nonlinear regression analysis of data points. The number of independent experiments (performed with different enzyme preparations on different days) is indicated by n.
| Activation of phosphorylation from ATP by cytoplasmic Ca2+, K0.5 | Activation of phosphorylation from Pi by luminal Ca2+, K0.5 | |
|---|---|---|
| μma | mmb | |
| S1a | 1.11 ± 0.02 (n = 9) | 7.27 ± 0.14 (n = 16) |
| S2a | 0.85 ± 0.04 (n = 2) | 8.39 ± 0.89 (n = 3) |
| S2b | 0.70 ± 0.04 (n = 3) | 4.83 ± 0.44 (n = 3) |
| S2b-T1032* | 0.75 ± 0.04 (n = 2) | 5.97 ± 0.33 (n = 3) |
| S2b-MFWS-AAAA | 0.88 ± 0.03 (n = 4) | 5.37 ± 0.29 (n = 3) |
| S1a2b | 0.59 ± 0.02 (n = 3) | 4.11 ± 0.28 (n = 3) |
| S1a2b-T1032* | 0.67 ± 0.02 (n = 2) | 4.21 ± 0.22 (n = 3) |
| S1a2b-L7/8 | 0.43 ± 0.02 (n = 2) | 3.24 ± 0.17 (n = 3) |
| S1a-L7/8 | 1.20 ± 0.04 (n = 2) | 5.74 ± 0.19 (n = 3) |
a Microsomes were pre-equilibrated at 0 °C in 40 mm MOPS/Tris (pH 7.0), 80 mm KCl, 5 mm MgCl2, 1 mm EGTA, and various concentrations of CaCl2 to obtain a series of free Ca2+ concentrations in the 0.1–100 μm range, followed by phosphorylation for 15 s at 0 °C with 5 μm [γ-32P]ATP and subsequent acid quenching. The Hill equation, EP = EPmax · [Ca2+]h/(K0.5h + [Ca2+]h ), was fitted to the data by nonlinear regression analysis (Hill coefficients (h) varying between 1.2 and 1.9).
b Data corresponding to Fig. 6.