Kinetics of cyanide and carbon monoxide dissociation from ferrous human haptoglobin:hemoglobin(II) complexes

Haptoglobin (Hp) counterbalances the adverse effects of extra-erythrocytic hemoglobin (Hb) trapping the αβ dimers of Hb. In turn, the Hp:Hb complexes display heme-based reactivity. Here, the kinetics of cyanide and carbon monoxide dissociation from ferrous-ligated Hp:Hb complexes are reported at pH 7.0 and 20.0 °C. Cyanide dissociation from Hp1-1:Hb(II)-CN^- and Hp2-2:Hb-CN^- has been followed upon the dithionite-mediated conversion of ferric to ferrous-ligated Hp:Hb complexes. Values of k_on for the dithionite-mediated reduction of Hp1-1:Hb(III)-CN^- and Hp2-2:Hb(III)-CN^- are (7.3 ± 1.1) × 10⁶ M^-1 s^-1 and (6.2 ± 1.0) × 10⁶ M^-1 s^-1, respectively. Values of the first-order rate constant (i.e., h) for cyanide dissociation from Hp1-1:Hb(II)-CN^- and Hp2-2:Hb(II)-CN^- are (1.2 ± 0.2) × 10^-1 s^-1 and (1.3 ± 0.2) × 10^-1 s^-1, respectively. CO dissociation from Hp:Hb(II)-CO complexes has been followed by replacing CO with NO. Values of the first-order rate constant (i.e., l) for CO dissociation from Hp1-1:Hb(II)-CO are (1.4 ± 0.2) × 10^-2 s^-1 and (6.2 ± 0.8) × 10^-3 s^-1, and those from Hp2-2:Hb(II)-CO are (1.3 ± 0.2) × 10^-2 s^-1 and (7.3 ± 0.9) × 10^-3 s^-1. Values of k_on, h, and l correspond to those reported for the R-state of tetrameric Hb and isolated α and β chains. This highlights the view that the conformation of the Hb αβ-dimers bound to Hp1-1 and Hp2-2 matches that of the R-state of the Hb tetramer. Furthermore, unlike ferric Hb(III), ligated ferrous Hb(II) does not show an assembly-linked structural change.