Monomer - Dimer equilibrium and oxygen binding properties of ferrous Vitreoscilla hemoglobin

L. Giangiacomo, M. Mattu, A. Arcovito, G. Bellenchi, M. Bolognesi, P. Ascenzi, A. Boffi

Research output: Contribution to journalArticle

Abstract

The monomer-dimer equilibrium and the oxygen binding properties of ferrous recombinant Vitreoscilla hemoglobin (Vitreoscilla Hb) have been investigated. Sedimentation equilibrium data indicate that the ferrous deoxygenated and carbonylated derivatives display low values of equilibrium dimerization constants, 6 × 102 and 1 × 102 M-1, respectively, at pH 7.0 and 10 °C. The behavior of the oxygenated species, as measured in sedimentation velocity experiments, is superimposable to that of the carbonylated derivative. The kinetics of O2 combination, measured by laser photolysis at pH 7.0 and 20 °C, is characterized by a second-order rate constant of 2 × 108 M-1 s-1 whereas the kinetics of O2 release at pH 7.0 is biphasic between 10 and 40 °C, becoming essentially monophasic below 10 °C. Values of the first-order rate constants (at 20 °C) and of the activation energies for the fast and slow phases of the Vitreoscilla Hb deoxygenation process are 4.2 s-1 and 19.2 kcal mol-1 and 0.15 s-1 and 24.8 kcal mol-1, respectively. Thus the biphasic kinetics of Vitreoscilla Hb deoxygenation is unrelated to the association state of the protein. The observed biphasic oxygen release may be accounted for by the presence of two different conformers in thermal equilibrium within the monomer. The two conformers may be assigned to a structure in which the heme-iron-bound ligand is stabilized by direct hydrogen bonding to TyrB 10 and a structure in which such interaction is absent. The slow interconversion between the two conformers may reflect a very large conformational rearrangement in the disordered distal pocket segment connecting helices C and E.

Original languageEnglish
Pages (from-to)9311-9316
Number of pages6
JournalBiochemistry
Volume40
Issue number31
DOIs
Publication statusPublished - Aug 7 2001

ASJC Scopus subject areas

  • Biochemistry

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    Giangiacomo, L., Mattu, M., Arcovito, A., Bellenchi, G., Bolognesi, M., Ascenzi, P., & Boffi, A. (2001). Monomer - Dimer equilibrium and oxygen binding properties of ferrous Vitreoscilla hemoglobin. Biochemistry, 40(31), 9311-9316. https://doi.org/10.1021/bi0101143