Fig 4. Mg²⁺ binds sites near the middle of the transmembrane domain and between the cytosolic A domain and N and P subdomains.

a, Side view of the overall dimeric structure with the left monomer in gray and the right monomer colored as in Fig. 1 with close-up views of the resolved Mg²⁺ ions and the nearby residues and proximal resolved water molecules (small red spheres). Site I is located within the TM α-helices 5, 7, and 8. Site II is comprised of residues from the A domain and P subdomain. Site III is comprised of residues from the A domain and N subdomain. b, MgtA_E331A (located in TM4) and MgtA_D780A (located in TM7) are unable to complement a Mg²⁺-auxotrophic E. coli strain. c, MgtA_D441A (site III) is only partially able to complement the Mg²⁺-auxotrophic E. coli strain upon overexpression. Complementation assays in b and c were carried out as for Fig. 3d (also see Extended Data Fig. 23).