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- Title
Reductive nitrosylation of ferric human hemoglobin bound to human haptoglobin 1-1 and 2-2.
- Authors
Ascenzi, Paolo; De Simone, Giovanna; Polticelli, Fabio; Gioia, Magda; Coletta, Massimo
- Abstract
Haptoglobin (Hp) sequesters hemoglobin (Hb) preventing the Hb-based damage occurring upon its physiological release into plasma. Here, reductive nitrosylation of ferric human hemoglobin [Hb(III)] bound to human haptoglobin (Hp) 1-1 and 2-2 [Hp1-1:Hb(III) and Hp2-2:Hb(III), respectively] has been investigated between pH 7.5 and 9.5, at <italic>T</italic>=20.0 °C. Over the whole pH range explored, only one process is detected reflecting NO binding to Hp1-1:Hb(III) and Hp2-2:Hb(III). Values of the pseudo-first-order rate constant for Hp1-1:Hb(III) and Hp2-2:Hb(III) nitrosylation (<italic>k</italic>) do not depend linearly on the ligand concentration but tend to level off. The conversion of Hp1-1:Hb(III)-NO to Hp1-1:Hb(II)-NO and of Hp2-2:Hb(III)-NO to Hp2-2:Hb(II)-NO is limited by the OH−- and H2O-based catalysis. In fact, bimolecular NO binding to Hp1-1:Hb(III), Hp2-2:Hb(III), Hp1-1:Hb(II), and Hp2-2:Hb(II) proceeds very rapidly. The analysis of data allowed to determine the values of the dissociation equilibrium constant for Hp1-1:Hb(III) and Hp2-2:Hb(III) nitrosylation [<italic>K</italic> = (1.2 ± 0.1) × 10−4 M], which is pH-independent, and of the first-order rate constant for Hp1-1:Hb(III) and Hp2-2:Hb(III) conversion to Hp1-1:Hb(II)-NO and Hp2-2:Hb(II)-NO, respectively (<italic>k</italic>′). From the dependence of <italic>k</italic>′ on [OH−], values of <italic>h</italic>OH- [(4.9 ± 0.6) × 103 M−1 s−1 and (6.79 ± 0.7) × 103 M−1 s−1, respectively] and of hH2O<inline-graphic></inline-graphic> [(2.6 ± 0.3) × 10−3 s−1] were determined. Values of kinetic and thermodynamic parameters for Hp1-1:Hb(III) and Hp2-2:Hb(III) reductive nitrosylation match well with those of the Hb R-state, which is typical of the αβ dimers of Hb bound to Hp.
- Subjects
HAPTOGLOBINS; NITROSYLATION; HEMOGLOBINS; BLOOD plasma; LIGAND binding (Biochemistry); THERMODYNAMICS
- Publication
Journal of Biological Inorganic Chemistry (JBIC), 2018, Vol 23, Issue 3, p437
- ISSN
0949-8257
- Publication type
Article
- DOI
10.1007/s00775-018-1551-y