Fig. 6. Liposome-based assay shows direct Mg²⁺ transport by CorB proteins.

a Liposomal assay showing direct transport of Mg²⁺ by MtCorB and TtCorB. Proteoliposomes (protein/lipid ratio of 1:30) or empty liposomes containing mag-fura-2 were equilibrated for 1 min before the addition of 5 mM MgCl₂ to initiate Mg²⁺ uptake (arrowhead). The data points represent mean ± SEM (n = 3 independent measurements). b An inward negative membrane potential generated by valinomycin leads to enhanced Mg²⁺ transport by TmCorA (protein/lipid ratio of 1:270) but not TtCorB (protein/lipid ratio of 1:30), suggesting electroneutral transport of Mg²⁺ by TtCorB. The data points represent mean ± SEM (n = 3 independent measurements). c Homology model of TtCorB showing locations of conserved residues in the negatively charged cavity and Mg²⁺ binding site. d, e Liposomal Mg²⁺ transport assays of TtCorB mutants under the same experimental conditions as panel (a). Mutations in the Mg²⁺-binding site increased transport activity while mutations near the π-helical turn reduced transport. The data points represent mean ± SEM (n = 3 independent measurements).