Ferric ion (hydr)oxo clusters in the “Venus flytrap” cleft of FbpA: Mössbauer, calorimetric and mass spectrometric studies

  • Arindam Mukherjee
  • Paul R. Bilton
  • Logan Mackay
  • Adam Janoschka
  • Haizhong Zhu
  • Dean Rea
  • Pat R. R. Langridge-Smith
  • Dominic J. Campopiano
  • Thomas Teschner
  • Alfred X. Trautwein
  • Volker Schünemann
  • Peter J. SadlerEmail author
Original Paper


Isothermal calorimetric studies of the binding of iron(III) citrate to ferric ion binding protein from Neisseria gonorrhoeae suggested the complexation of a tetranuclear iron(III) cluster as a single step binding event (apparent binding constant K app ITC  = 6.0(5) × 105 M−1). High-resolution Fourier transform ion cyclotron resonance mass spectrometric data supported the binding of a tetranuclear oxo(hydroxo) iron(III) cluster of formula [Fe4O2(OH)4(H2O)(cit)]+ in the interdomain binding cleft of FbpA. The mutant H9Y-nFbpA showed a twofold increase in the apparent binding constant [K app ITC  = 1.1(7) × 106 M−1] for the tetranuclear iron(III) cluster compared to the wild-type protein. Mössbauer spectra of Escherichia coli cells overexpressing FbpA and cultured in the presence of added 57Fe citrate were indicative of the presence of dinuclear and polynuclear clusters. FbpA therefore appears to have a strong affinity for iron clusters in iron-rich environments, a property which might endow the protein with new biological functions.

Graphical Abstract


Mössbauer spectroscopy Ferric ion binding protein A Bacterial transferrin Isothermal titration calorimetry Iron clusters Fourier transform mass spectrometry 



Electrospray ionization mass spectrometry


Ferric ion binding protein A


Ferric ion binding protein B


Ferric ion binding protein C


Ferric ion binding protein A from Haemophilus influenzae


Ferric ion binding protein A from Neisseria gonorrhoeae


Fourier transform ion cyclotron resonance


Inductively coupled plasma mass spectrometry


Isothermal titration calorimetry


Phosphate transport system protein



We thank the European Commission for a Marie Curie Fellowship for A.M., the BBSRC (RASOR, IRColl in Proteomic Technologies), EPSRC (WCAS), AWM/ERDF (Science City) and ERC (grant no. 247450, BIOINCMED) for funding, the Universities of Edinburgh and Warwick for analytical facilities, and Dr Teresa Pinheiro for access to isothermal calorimetry. A.M. thanks IISER Kolkata for financial support. V.S. acknowledges the federal state Rheinland-Pfalz of Germany for financial support within the framework of NANOKAT.

Supplementary material

775_2012_878_MOESM1_ESM.pdf (239 kb)
Supplementary material 1 (PDF 238 kb)


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Copyright information

© SBIC 2012

Authors and Affiliations

  • Arindam Mukherjee
    • 1
    • 2
  • Paul R. Bilton
    • 3
  • Logan Mackay
    • 4
  • Adam Janoschka
    • 5
  • Haizhong Zhu
    • 3
  • Dean Rea
    • 2
  • Pat R. R. Langridge-Smith
    • 4
  • Dominic J. Campopiano
    • 3
  • Thomas Teschner
    • 6
  • Alfred X. Trautwein
    • 6
  • Volker Schünemann
    • 5
  • Peter J. Sadler
    • 2
    Email author
  1. 1.Department of Chemical SciencesIndian Institute of Science Education and Research KolkataNadiaIndia
  2. 2.Department of ChemistryUniversity of WarwickCoventryUK
  3. 3.School of ChemistryUniversity of EdinburghEdinburghUK
  4. 4.SIRCAMS, School of ChemistryUniversity of EdinburghEdinburghUK
  5. 5.University of KaiserslauternKaiserslauternGermany
  6. 6.University of LübeckLübeckGermany

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