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Issue 2, 2017
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Magnetic circular dichroism studies of iron(II) binding to human calprotectin

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Abstract

Calprotectin (CP) is an abundant metal-chelating protein involved in host defense, and the ability of human CP to bind Fe(II) in a calcium-dependent manner was recently discovered. In the present study, near-infrared magnetic circular dichroism spectroscopy is employed to investigate the nature of Fe(II) coordination at the two transition-metal-binding sites of CP that are a His3Asp motif (site 1) and a His6 motif (site 2). Upon the addition of sub-stoichiometric Fe(II), a six-coordinate (6C) Fe(II) center associated with site 2 is preferentially formed in the presence of excess Ca(II). This site exhibits an exceptionally large ligand field (10Dq = 11 045 cm−1) for a non-heme Fe(II) protein. Analysis of CP variants lacking residues of the His6 motif supports that CP coordinates Fe(II) at site 2 by employing six His ligands. In the presence of greater than one equiv. of Fe(II) or upon mutation of the His6 motif, the metal ion also binds at site 1 of CP to form a five-coordinate (5C) Fe(II)–His3Asp motif that was previously unidentified in this system. Notably, the introduction of His-to-Ala mutations at the His6 motif results in a mixture of 6C (site 2) and 5C (site 1) signals in the presence of sub-stoichiometric Fe(II). These results are consistent with a reduced Fe(II)-binding affinity of site 2 as more weakly coordinating water-derived ligands complete the 6C site. In the absence of Ca(II), both sites 1 and 2 are occupied upon addition of sub-stoichiometric Fe(II), and a stronger ligand field is observed for the 5C site. These spectroscopic studies provide further evaluation of a unique non-heme Fe(II)–His6 site for metalloproteins and support the notion that Ca(II) ions influence the Fe(II)-binding properties of CP.

Graphical abstract: Magnetic circular dichroism studies of iron(ii) binding to human calprotectin

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Publication details

The article was received on 04 Aug 2016, accepted on 11 Oct 2016 and first published on 11 Oct 2016


Article type: Edge Article
DOI: 10.1039/C6SC03487J
Citation: Chem. Sci., 2017,8, 1369-1377
  • Open access: Creative Commons BY-NC license
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    Magnetic circular dichroism studies of iron(II) binding to human calprotectin

    T. M. Baker, T. G. Nakashige, E. M. Nolan and M. L. Neidig, Chem. Sci., 2017, 8, 1369
    DOI: 10.1039/C6SC03487J

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