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Relaxometric characterization of human hemalbumin

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Abstract

Hemalbumin [i.e., Fe(III)-protoporphyrin IX-human serum albumin; Fe(III)heme-HSA] is an important intermediate in the recovery of heme iron following hemolysis. Relaxometric data are consistent with the occurrence of a hexacoordinated high-spin Fe(III) center with no water in the inner coordination sphere. The relatively high relaxation enhancement observed for an aqueous solution of Fe(III)heme-HSA (r 1p=4.8 mM–1 s–1 at 20 MHz, pH 7, and 25 °C) is ascribed to the occurrence of a strong contribution from water molecules in the second coordination sphere. Structural analysis of the putative binding region has been performed by a Monte Carlo simulated annealing procedure, which allowed us to identify His105 and Tyr148 as axial ligands. The role of a tyrosinate as the sixth Fe(III)heme ligand is supported by the pH-dependent analysis. Interestingly, when Fe(III) is replaced by Mn(III), the occurrence of a fast exchanging water molecule at pH values close to neutrality is detected. As the pH is increased, the Mn(III) containing system behaves analogously to Fe(III)heme-HSA. At higher pH, the phenolate ligand is eventually displaced by OH from both Fe(III) and Mn(III) centers. Support for the proposed bonding scheme has been gained also from competitive binding assays for the sixth coordination site by fluoride, azide, and imidazole ligands.

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Abbreviations

BSA:

bovine serum albumin

Fe(III)heme-HSA:

Fe(III)-protoporphyrin IX-HSA complex

FID:

free induction decay

Hb:

hemoglobin

HSA:

human serum albumin

Hb:

myoglobin

Mn(III)heme-HSA:

Mn(III)-protoporphyrin IX-HSA

NMRD:

nuclear magnetic relaxation dispersion

PRE:

proton relaxation enhancement

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Correspondence to Silvio Aime.

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Fasano, M., Baroni, S., Vannini, A. et al. Relaxometric characterization of human hemalbumin. JBIC 6, 650–658 (2001). https://doi.org/10.1007/s007750100242

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  • DOI: https://doi.org/10.1007/s007750100242

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