Oxygen equilibrium studies of cross-linked asymmetrical cyanomet valency hybrid hemoglobins: models for partially oxygenated species.

Abstract

Oxygen equilibrium curves have been measured to determine the binding constant at each oxygenation step (Ki) for various cross-linked hemoglobins, (alpha beta)A(alpha beta)CXL, (alpha +CN beta)A(alpha beta)CXL, (alpha beta +CN)A(alpha beta)CXL, (alpha +CN beta +CN)A(alpha beta)CXL, and (alpha beta +CN)A(alpha +CN beta)CXL, where the subscripts A and C denote that the alpha beta dimer is derived from human normal adult hemoglobin and mutant hemoglobin C (beta 6Glu----Lys), respectively, and XL denotes cross-linking between the lysyl residues at position 82 in the two beta chains by bis(3,5-dibromosalicyl) fumarate as described by Miura and Ho [Miura, S., & Ho, C. (1982) Biochemistry 21, 6280-6287]. The oxygen equilibrium data indicate that the oxygen affinity increases with the number of cyanomet hemes carried by the cross-linked mixed valency hybrid hemoglobins. The oxygen binding property depends not only on the number of the subunits carrying cyanomet hemes but also on the distribution of cyanomet hemes among the four subunits. A striking effect is observed in singly cyanomet valency hybrid hemoglobins; namely, (alpha +CN beta)A(alpha beta)CXL exhibits lower oxygen affinity and higher cooperativity than (alpha beta +CN)A(alpha beta)CXL. The magnitude of the Adair constants and their pH dependency of (alpha +CN beta)A(alpha beta)CXL (Ki, i = 1-3) are analogous to those of the Adair constants of (alpha beta)A(alpha beta)CXL (Ki, i = 2-4), whereas such an analogy is not observed between (alpha beta +CN)A(alpha beta)CXL and (alpha beta)A(alpha beta)CXL. The doubly cyanomet mixed valency hybrid cross-linked hemoglobins exhibit high oxygen affinity and reduced cooperativity, and their Adair constants are not analogous to K3 and K4 of (alpha beta)A(alpha beta)CXL.(ABSTRACT TRUNCATED AT 250 WORDS)