All extant life forms require trace transition metals (e.g., Fe2/3+, Cu1/2+, and Mn2+) to survive. However, as these are environmentally scarce, organisms have evolved sophisticated metal uptake machineries. In bacteria, high-affinity import of transition metals is predominantly mediated by ABC transporters. During bacterial infection, sequestration of metal by the host further limits the availability of these ions, and accordingly, bacterial ABC transporters (importers) of metals are key virulence determinants. However, the structure-function relationships of these metal transporters have not been fully elucidated. Here, we used metal-sensitivity assays, advanced structural modeling, and enzymatic assays to study the ABC transporter MntBC-A, a virulence determinant of the bacterial human pathogen Bacillus anthracis. We find that despite its broad metal-recognition profile, MntBC-A imports only manganese, whereas zinc can function as a high-affinity inhibitor of MntBC-A. Computational analysis shows that the transmembrane metal permeation pathway is lined with six titratable residues that can coordinate the positively charged metal, and mutagenesis studies show that they are essential for manganese transport. Modeling suggests that access to these titratable residues is blocked by a ladder of hydrophobic residues, and ATP-driven conformational changes open and close this hydrophobic seal to permit metal binding and release. The conservation of this arrangement of titratable and hydrophobic residues among ABC transporters of transition metals suggests a common mechanism. These findings advance our understanding of transmembrane metal recognition and permeation and may aid the design and development of novel antibacterial agents.
Keywords: ABC transporter; ATPase; membrane proteins; metal binding; transition metals; virulence.
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