Oxygen Equilibrium of Hemoglobins Containing Unnatural Hemes

EFFECT OF MODIFICATION OF HEME CARBOXYL GROUPS AND SIDE CHAINS AT POSITIONS 2 AND 4

From the ¹ From the Department of Biochemistry, Kanazawa University School of Medicine, Kanazawa, Japan

Stable ferro-forms of hemoglobins containing various hemes such as dimethyl proto-, dimethyl meso-, etio-, meso-, hemato-, and deuteroheme were prepared by combination of human globin with hemes, followed by reduction with dithionite and purification with carboxymethyl Sephadex column. Sedimentation coefficients of these hemoglobins were all near to 4.0 S and chemical analysis showed that 4 moles of heme were bound per mole of hemoglobin.

Esterification of propionyl carboxyl groups of heme did not change the absorption spectra of hemoglobin; absorption spectra of dimethyl protohemoglobin were essentially identical with those of protohemoglobin, and absorption spectra of dimethyl mesohemoglobin with those of mesohemoglobin. Absorption spectra of etiohemoglobin, in which the propionic acid groups of mesoheme were substituted by ethyl groups, were similar to those of mesohemoglobin. Parameters of oxygen equilibrium, namely oxygen affinity, n in Hill's equation, and Bohr effect, of hemoglobin containing hemes which lack free carboxyl groups were the same as those for the corresponding hemoglobin with free propionic acid groups.

Oxygen affinities of meso-, deutero-, hemato-, and protohemoglobin, in which heme side chains at positions 2 and 4 are ethyl, hydrogen, hydroxyethyl, and vinyl groups, respectively, were in the ratio of 5:2:1.3:1, corresponding to the reverse order of the negative inductive effects of the groups at positions 2 and 4. These results suggest the effect of electron of a porphyrin ring on the oxygen binding.

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