Properties of hemoglobin and dextran-hemoglobin rightshifted by oxidized inositol tetrakisphosphate

Phytate was digested by wheat bran phytase to yield inositol tetrakisphosphate. Periodate-oxidized inositol tetrakisphosphate (oxyIP4) was coupled by means of reductive alkylation to hemoglobin and the covalent dextran-hemoglobin conjugate to yield the rightshifted (rs) compounds rsHb and rsDxHb, respectively. The variations of the oxygen dissociation curves of these molecules with pH and temperature were compared to those of hemoglobin. The variations with pH were found to be less pronounced for these rightshifted forms. An extensive decrease in the half-saturation oxygen tension was observed, however, with both rsHb and rsDxHb, as in the case of unmodified Hb. Modification of hemoglobin by oxyIP4 at the polyphosphate site was suggested by the lack of a further rightshifting effect of phytate on rsHb, and by the similarity between the difference spectrum of rs-methemoglobin and the difference spectrum induced by the addition of phytate.