Proton-linked subunit heterogeneity in ferrous nitrosylated human adult hemoglobin: An EPR study

Paolo Ascenzi, Alessio Bocedi, Mauro Fasano, Magda Gioia, Stefano Marini, Massimo Coletta

Research output: Contribution to journalArticlepeer-review

Abstract

The effect of pH on the X-band electron paramagnetic resonance (EPR) spectrum of ferrous nitrosylated human adult tetrameric hemoglobin (HbNO) as well as of ferrous nitrosylated monomeric α- and β-chains has been investigated, at -163°C. At pH 7.3, the X-band EPR spectrum of tetrameric HbNO and ferrous nitrosylated monomeric α- and β-chains displays a rhombic shape. Lowering the pH from 7.3 to 3.0, tetrameric HbNO and ferrous nitrosylated monomeric α- and β-chains undergo a transition towards a species characterized by a X-band EPR spectrum with a three-line splitting centered at 334 mT. These pH-dependent spectroscopic changes may be taken as indicative of the cleavage, or the severe weakening, of the proximal HisF8-Fe bond. In tetrameric HbNO, the pH-dependent spectroscopic changes depend on the acid-base equilibrium of two apparent ionizing groups with pKa values of 5.8 and 3.8. By contrast, the pH-dependent spectroscopic changes occurring in ferrous nitrosylated monomeric α- and β-chains depend on the acid-base equilibrium of one apparent ionizing group with pKa values of 4.8 and 4.7, respectively. The different pKa values for the proton-linked spectroscopic transition(s) of tetrameric HbNO and ferrous nitrosylated monomeric α- and β-chains suggest that the quaternary assembly drastically affects the strength of the proximal HisF8-Fe bond in both subunits. This probably reflects a 'quaternary effect', i.e., structural changes in both subunits upon tetrameric assembly, which is associated to a relevant variation of functional properties (i.e., proton affinity).

Original languageEnglish
Pages (from-to)1255-1259
Number of pages5
JournalJournal of Inorganic Biochemistry
Volume99
Issue number5
DOIs
Publication statusPublished - May 2005

Keywords

  • EPR spectroscopy
  • Ferrous nitrosylated monomeric α- and β-chains
  • Ferrous nitrosylated tetrameric human adult hemoglobin
  • Proton-linked subunit heterogeneity in ferrous nitrosylated human adult tetrameric hemoglobin

ASJC Scopus subject areas

  • Biochemistry
  • Inorganic Chemistry

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