Studies on cobalt myoglobins and hemoglobins. XI. The interaction of carbon monoxide and oxygen with α and β subunits in iron-cobalt hybrid hemoglobins

Masao Ikeda-Saito, Takashi Yonetani

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28 Citations (Scopus)

Abstract

An artificial hybrid hemoglobin, α(Co)2β(Fe)2, the α and β subunits of which carry cobaltous protoporphyrin IX and ferrous protoporphyrin IX, respectively, and its complementary hybrid α(Fe)2β(Co)2 were prepared and the properties of their ferrous subunits were examined by equilibrium and kinetic measurements with carbon monoxide as a ligand. The β(Fe)2 subunits in fully deoxy α(Co)2β(Fe)2 exhibited a higher affinity for carbon monoxide than did the α(Fe)2 subunits in fully deoxy α(Fe)2β(Co)2. Addition of 2 mm-inositol hexaphosphate decreased fourfold the carbon monoxide affinity of the α(Fe)2 subunits in α(Fe)2β(Co)2 and by more than tenfold that of the β(Fe)2 subunits in α(Co)2β(Fe)2 at pH 7.0. The higher affinity for carbon monoxide of the β(Fe)2 subunits inα(Co)2β(Fe)2 than that of the α(Fe)2 subunits in α(Fe)2β(Co)2 was caused by a smaller dissociation rate of the β(Fe)2-carbon monoxide complex. These results are considered to underlie the different affinity for carbon monoxide of the α and β subunits in deoxy hemoglobin. Oxygen equilibria of the cobaltous subunits in iron-cobalt hybrid hemoglobins were also measured in the presence of carbon monoxide. The α(Co)2 subunits in α(Co)2β(FeCO)2 showed a higher oxygen affinity than the β(Co)2 subunits in α(FeCO)2β(Co)2. Inositol hexaphosphate lowered the oxygen affinity of the β(Co)2 subunits in α(FeCO)2β(Co)2 by eight-fold, but that of the α(Co)2 subunits in α(Co)2β(FeCO)2|by only 1.6-fold. The magnitude of the alkaline Bohr effect, as defined by Δlog Pm ΔpH was found to be -0.34 and -0.14 for α(FeCO)2β(Co)2 and α(Co)2β(FeCO)2, respectively, in 0.1 m-phosphate buffer at 15 °C. The rate of oxygenation and deoxygenation of the cobaltous subunits in iron-cobalt hybrid hemoglobins in the presence of carbon monoxide was determined by a temperature-jump relaxation method in 0.1 m-phosphate buffer with and without inositol hexaphosphate. Their relaxation spectra were of a single exponential character and differed from that of cobalt hemoglobin. Without inositol hexaphosphate, the association rate constants for both α(Co)2β(FeCO)2 and α(FeCO)2β(Co)2 were close to that for cobalt hemoglobin, whereas the dissociation rate constants for iron-cobalt hybrid hemoglobins were smaller than that for cobalt hemoglobin by more than fourfold. Inositol hexaphosphate affected both the association and dissociation rates of α(FeCO)2β(Co)2 but did so to a lesser extent than those of α(Co)2β(FeCO)2. These observations suggest strongly a different role for the α and β subunits in the co-operative oxygenation and alkaline Bohr effect of hemoglobin.

Original languageEnglish
Pages (from-to)845-858
Number of pages14
JournalJournal of Molecular Biology
Volume138
Issue number4
DOIs
Publication statusPublished - 1980 Apr 25

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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