Periplasmic metal-resistance protein CusF exhibits high affinity and specificity for both CuI and AgI.
The periplasmic protein CusF, as a part of the CusCFBA efflux complex, plays a role in resistance to elevated levels of copper and silver in Escherichia coli. Although homologues have been identified in other Gram-negative bacteria, the substrate of CusF and its precise role in metal resistance have not been described. Here, isothermal titration calorimetry (ITC) was used to demonstrate that CusF binds with high affinity to both Cu(I) and Ag(I) but not Cu(II). The affinity of CusF for Ag(I) was higher than that for Cu(I), which could reflect more efficient detoxification of Ag(I) given the lack of a cellular need for Ag(I). The chemical shifts in the nuclear magnetic resonance (NMR) spectra of CusF-Ag(I) as compared to apo-CusF show that the region of CusF most affected by Ag(I) binding encompasses three absolutely conserved residues: H36, M47, and M49. This suggests that these residues may play a role in Ag(I) coordination. The NMR spectra of CusF in the presence of Cu(II) do not indicate specific binding, which is in agreement with the ITC data. We conclude that Cu(I) and Ag(I) are the likely physiological substrates.
Kittleson
Joshua T
JT
Departments of Biochemistry and Molecular Biophysics, University of Arizona, Tucson, Arizona 85721, USA.
Loftin
Isabell R
IR
Hausrath
Andrew C
AC
Engelhardt
Kevin P
KP
Rensing
Christopher
C
McEvoy
Megan M
MM
D001665:Binding Sites; D002151:Calorimetry / Q000379:methods; D027682:Cation Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D003300:Copper / Q000737:chemistry / Q000378:metabolism*; D000079922:Copper Transport Proteins; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins; D007454:Iodides / Q000737:chemistry / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D011487:Protein Conformation; D018030:Silver Compounds / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity
D027682:Cation Transport Proteins; D000079922:Copper Transport Proteins; C506213:CusF protein, E coli; D029968:Escherichia coli Proteins; D007454:Iodides; D018030:Silver Compounds; D003300:Copper; C073870:cuprous iodide; C030584:silver iodide