REEVALUATION OF CHLORIDES REGULATION OF HEMOGLOBIN OXYGEN-UPTAKE - THE NEGLECTED CONTRIBUTION OF PROTEIN HYDRATION IN ALLOSTERISM

We have measured hemoglobin oxygen uptake vs. the partial pressure of oxygen, with independently controlled activities of chloride and water . This control is effected by combining different concentrations of Na Cl and sucrose in the bathing solution to achieve: (i) water activitie s were varied and CI- activity was fixed, (ii) both water and CI- acti vities were varied with a traditional NaCI titration, or (iii) CI- act ivities were varied and water activity was fixed by adding compensatin g sucrose. Within this analysis, the CI--regulated loading of four oxy gens can be described by the reaction Hb.CI- + 4 O-2 + 65 H2O reversib le arrow Hb.4O(2).65H(2)O + CI-. The dissociation of a neatly integral chloride, rather than the nonintegral 1.6 chlorides inferred earlier from simple salt titration, demonstrates the need to recognize the pot entially large contribution from changes in water activity when titrat ing weakly binding solutes. The single-chloride result might simplify structural considerations of the action of CI- in hemoglobin regulatio n.