Figure 4. Mg²⁺and pH dependency of MgtA.

(A) ATP hydrolysis was measured with increasing concentration Mg²⁺ at fixed ATP concentrations. Irrespective of the ATP concentration used, maximum ATPase activity was observed at around 13 μM P_i min^-1 mg^-1 and increasing Mg²⁺ concentration decreases the ATPase activity. (B) Data from [A] was plotted against Mg²⁺_free in assay condition determined using MAXC as mentioned in the methods section. (C) Data from [B] was plotted in the range from 0 to 120 μM Mg²⁺_free and used for kinetic calculations summarized in Table 1. (D) ATP hydrolysis was measured with increasing ATP concentration at various fixed Mg²⁺ concentrations. Since ATP acts as Mg²⁺ chelator, the Mg²⁺_free concentration decreases with increasing ATP concentration. (E) ATP hydrolysis measured at various fixed pH conditions while increasing the Mg²⁺ concentration. Bis-tris propane at indicated pH values was used as buffer. (F) Data from [E] was plotted against calculated Mg²⁺_free. A-F, Values plotted are mean ± SD (n = 3).

DOI: http://dx.doi.org/10.7554/eLife.11407.016

Figure 4—figure supplement 1. ATP dependency of MgtA at fixed Mg²⁺ concentration.

Solid line represents the ATPase activity of MgtA and dotted line represents the Mg²⁺_free available at corresponding ATP concentrations. Colour of the dotted line corresponds to the concentration of Mg²⁺_total used to calculate Mg²⁺_free. The graph shows that MgtA achieves maximum activity when the Mg²⁺_free concentration in the assay setup is below 50 μM. Values plotted are mean ± SD (n = 3).

DOI: http://dx.doi.org/10.7554/eLife.11407.018