Internal electron transfer between hemes and Cu(II) bound at cysteine β93 promotes methemoglobin reduction by carbon monoxide

Previous studies showed that CO/H2O oxidation provides electrons to drive the reduction of oxidized hemoglobin (metHb), We report here that Cu(II) addition accelerates the rate of metHb beta chain reduction by CO by a factor of about 1000, A mechanism whereby electron transfer occurs via an internal pathway coupling CO/H2O oxidation to Fe(III) and Cu(II) reduction is suggested by the observation that the copper-induced rate enhancement is inhibited by blocking Cys-beta 93 with N-ethylmaleimide. Furthermore, this internal electron-transfer pathway is more readily established at low Cu(II) concentrations in Hb Deer Lodge (beta 2His --> Arg) and other species lacking His-beta 2 than in Hb A(0). This difference is consistent with preferential binding of Cu(II) in Hb A(0) to a high affinity site involving His-beta 2, which is ineffective in promoting electron exchange between Cu(II) and the beta heme iron, Effective electron transfer is thus affected by Hb type but is not governed by the R <-> T conformational equilibrium. The beta hemes in Cu(II)-metHb are reduced under CO at rates close to those observed for cytochrome c oxidase, where heme and copper are present together in the oxygen-binding site and where internal electron transfer also occurs.