FIGURE 4.

Mg²⁺ requirement for DgkB binding to PtdGro vesicles. DgkB binding to phospholipid vesicles was measured using the AlphaScreen proximity assay that signals the association of DgkB with 100-nm diameter phospholipid vesicles. Assays were conducted in triplicate and the dissociation constants calculated (value ± S.E.). A, phospholipid vesicles were composed of PtdGro and binding assays were performed in the presence of the indicated concentration of MgCl₂. B, comparison of the binding of wild-type DgkB and three mutants in the aspartate residues that participate in the Mg²⁺·Asp·water network using the AlphaScreen assay in the presence of PtdGro vesicles and 10 mm Mg²⁺. The apparent K_d values are listed in Table 1. The symbols are: DgkB, •; DgkB[D216A], ▪; DgkB[D124A], ○; and DgkB[D271A], □. Error bars represent standard error.