title: Mechanism of ATPase-mediated Cu+ export and delivery to periplasmic chaperones: the interaction of Escherichia coli CopA and CusF.
abstract: Cellular copper homeostasis requires transmembrane transport and compartmental trafficking while maintaining the cell essentially free of uncomplexed Cu(2+/+). In bacteria, soluble cytoplasmic and periplasmic chaperones bind and deliver Cu(+) to target transporters or metalloenzymes. Transmembrane Cu(+)-ATPases couple the hydrolysis of ATP to the efflux of cytoplasmic Cu(+). Cytosolic Cu(+) chaperones (CopZ) interact with a structural platform in Cu(+)-ATPases (CopA) and deliver copper into the ion permeation path. CusF is a periplasmic Cu(+) chaperone that supplies Cu(+) to the CusCBA system for efflux to the extracellular milieu. In this report, using Escherichia coli CopA and CusF, direct Cu(+) transfer from the ATPase to the periplasmic chaperone was observed. This required the specific interaction of the Cu(+)-bound form of CopA with apo-CusF for subsequent metal transfer upon ATP hydrolysis. As expected, the reverse Cu(+) transfer from CusF to CopA was not observed. Mutation of CopA extracellular loops or the electropositive surface of CusF led to a decrease in Cu(+) transfer efficiency. On the other hand, mutation of Met and Glu residues proposed to be part of the metal exit site in the ATPase yielded enzymes with lower turnover rates, although Cu(+) transfer was minimally affected. These results show how soluble chaperones obtain Cu(+) from transmembrane transporters. Furthermore, by explaining the movement of Cu(+) from the cytoplasmic pool to the extracellular milieu, these data support a mechanism by which cytoplasmic Cu(+) can be precisely directed to periplasmic targets via specific transporter-chaperone interactions.
authors.affiliation: From the Department of Chemistry and Biochemistry, Worcester Polytechnic Institute, Worcester, Massachusetts 01609 and.
authors.forename: Teresita
authors.initials: T
authors.lastname: Padilla-Benavides
authors.affiliation: the Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721.
authors.forename: Alayna M
authors.initials: AM
authors.lastname: George Thompson
authors.affiliation: the Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721 mcevoy@arizona.edu.
authors.forename: Megan M
authors.initials: MM
authors.lastname: McEvoy
authors.affiliation: From the Department of Chemistry and Biochemistry, Worcester Polytechnic Institute, Worcester, Massachusetts 01609 and arguello@wpi.edu.
authors.forename: José M
authors.initials: JM
authors.lastname: Argüello
mesh_terms: D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000235:genetics / Q000378:metabolism; D027682:Cation Transport Proteins / Q000235:genetics / Q000378:metabolism*; D003300:Copper / Q000378:metabolism*; D000079922:Copper Transport Proteins; D000073840:Copper-Transporting ATPases; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D017136:Ion Transport / Q000502:physiology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D033903:Periplasmic Proteins / Q000235:genetics / Q000378:metabolism*; D017433:Protein Structure, Secondary
chemical_list: D027682:Cation Transport Proteins; D000079922:Copper Transport Proteins; C096350:CusC protein, E coli; C506213:CusF protein, E coli; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D033903:Periplasmic Proteins; D003300:Copper; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D000073840:Copper-Transporting ATPases; C000612460:copA protein, E coli
keywords: Chaperone; Copper; Copper ATPase; Copper Transport; Metal Homeostasis; Metal Ion-Protein Interaction