JBIC Journal of Biological Inorganic Chemistry

, Volume 13, Issue 2, pp 271–288

Structure and metal binding properties of ZnuA, a periplasmic zinc transporter from Escherichia coli

  • Liliya A. Yatsunyk
  • J. Allen Easton
  • Lydia R. Kim
  • Stacy A. Sugarbaker
  • Brian Bennett
  • Robert M. Breece
  • Ivan I. Vorontsov
  • David L. Tierney
  • Michael W. Crowder
  • Amy C. Rosenzweig
Original Paper

Abstract

ZnuA is the periplasmic Zn2+-binding protein associated with the high-affinity ATP-binding cassette ZnuABC transporter from Escherichia coli. Although several structures of ZnuA and its homologs have been determined, details regarding metal ion stoichiometry, affinity, and specificity as well as the mechanism of metal uptake and transfer remain unclear. The crystal structures of E. coli ZnuA (Eco-ZnuA) in the apo, Zn2+-bound, and Co2+-bound forms have been determined. ZnZnuA binds at least two metal ions. The first, observed previously in other structures, is coordinated tetrahedrally by Glu59, His60, His143, and His207. Replacement of Zn2+ with Co2+ results in almost identical coordination geometry at this site. The second metal binding site involves His224 and several yet to be identified residues from the His-rich loop that is unique to Zn2+ periplasmic metal binding receptors. Electron paramagnetic resonance and X-ray absorption spectroscopic data on CoZnuA provide additional insight into possible residues involved in this second site. The second site is also detected by metal analysis and circular dichroism (CD) titrations. Eco-ZnuA binds Zn2+ (estimated Kd < 20 nM), Co2+, Ni2+, Cu2+, Cu+, and Cd2+, but not Mn2+. Finally, conformational changes upon metal binding observed in the crystal structures together with fluorescence and CD data indicate that only Zn2+ substantially stabilizes ZnuA and might facilitate recognition of ZnuB and subsequent metal transfer.

Keywords

ZnuA Zinc-specific uptake system Zinc binding ATP binding cassette transporter 

Abbreviations

ABC

ATP-binding cassette

ANS

8-Anilino-1-naphthalenesulfonic acid

apoZnuA

Apo form of ZnuA protein from Escherichia coli

CD

Circular dichroism

CoZnuA

Co2+-loaded form of ZnuA protein from Escherichia coli

Eco-ZnuA

ZnuA protein from Escherichia coli

EPR

Electron paramagnetic resonance

EXAFS

Extended X-ray absorption fine structure

HEPES

N-(2-Hydroxyethyl)piperazine-N′-ethanesulfonic acid

Hin-Pzp1

Zn2+-binding protein from Haemophilus influenzae

ICP-AES

Inductively coupled plasma atomic emission spectroscopy

MBR

Metal-binding receptor

MF

Mag-Fura-2

MOPS

3-(N-Morpholino)propanesulfonic acid

PDB

Protein Data Bank

PEG

Poly(ethylene glycol)

PLBP

Periplasmic ligand-binding proteins

RMSD

RMS deviation

Spn-PsaA

Mn2+-binding protein from Streptococcus pneumoniae

Syn-MntC

Mn2+-binding protein from Synechocystis PCC sp. 6803

Syn-ZnuA

Zn2+-binding protein from Synechocystis PCC sp. 6803

Tpa-TroA

Mn2+-binding protein from Treponema pallidum

Tris

Tris(hydroxymethly)aminomethane

Znu

Zn2+-specific uptake system

ZnZnuA

Zn2+-loaded form of ZnuA protein from Escherichia coli

Supplementary material

775_2007_320_MOESM1_ESM.pdf (1.7 mb)
ESM1 (PDF 1.65 MB)

Copyright information

© SBIC 2007

Authors and Affiliations

  • Liliya A. Yatsunyk
    • 1
  • J. Allen Easton
    • 2
  • Lydia R. Kim
    • 1
  • Stacy A. Sugarbaker
    • 2
  • Brian Bennett
    • 3
  • Robert M. Breece
    • 4
  • Ivan I. Vorontsov
    • 5
  • David L. Tierney
    • 4
  • Michael W. Crowder
    • 2
  • Amy C. Rosenzweig
    • 1
  1. 1.Departments of Biochemistry, Molecular Biology, and Cell Biology and of ChemistryNorthwestern UniversityEvanstonUSA
  2. 2.Department of Chemistry and BiochemistryMiami UniversityOxfordUSA
  3. 3.National Biomedical EPR Center, Department of BiophysicsMedical College of WisconsinMilwaukeeUSA
  4. 4.Department of Chemistry and Chemical BiologyUniversity of New MexicoAlbuquerqueUSA
  5. 5.Department of Molecular Pharmacology and Biological ChemistryNorthwestern University Feinberg School of MedicineChicagoUSA

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