A POLYMERIZING ROOT-EFFECT FISH HEMOGLOBIN WITH HIGH SUBUNIT HETEROGENEITY - CORRELATION WITH PRIMARY STRUCTURE

The blood of the teleost Chelodonichthys kumu, living in the temperate waters of New Zealand, contains a single hemoglobin. The complete ami no acid sequence of the alpha and beta chain has been established. The presence of a reactive Cys in the external position betaCD8(49) cause s polymerisation through intermolecular disulfide bridges between beta chains, with no alteration of functional features. C. kumu Root-effec t hemoglobin displays very low or no subunit co-operativity in the phy siological pH range. Kinetic experiments on the oxygen dissociation an d binding of carbon monoxide show a marked, pH-dependent functional he terogeneity of the two chains, which contributes to the observed reduc tion of co-operativity. In contrast, kinetic heterogeneity was not obs erved in the process of CO dissociation, indicating that functional di fferences between the subunits are detectable only for the dynamic lig and association pathway. The allosteric effector, ATP, seems to increa se the pK(a) of the proton-linked effect on the slow-reacting subunit, affecting the quaternary equilibrium through stabilisation of the T s tate at lower pH, rather than enhancing the functional heterogeneity i tself In position E11 of both chains, Val (usually present at the dist al side of the heme), is substituted by Ile. Although this residue has been shown not to significantly alter ligand binding to the a chain, to some extent it can perturb the access of oxygen to the beta chain. Thus, this substitution may be the main reason for subunit functional heterogeneity.