Direct electrochemistry of engineered cytochrome b562 molecules with a ligand binding pocket

Abstract

The rapid and reversible electron transfer reaction of cytochrome b 562 was observed at an In 2O 3 electrode. The estimated heterogeneous electron transfer rate constant ( k 0′) was k 0′ ≥ 5.0 × 10 −3 cm s −1 at pH 6.5. When the methionine-7 (Met-7) residue, which coordinates to the heme iron as an axial ligand, of the wild-type cytochrome b 562 was replaced by an Ala or Gly residue, a water molecule bound to the heme iron and the electron transfer rate constants decreased to 1.3 × 10 −3 and 1.8 × 10 −3 cm s −1, respectively. This decrease in the electron transfer rate would be due to the larger reorganization energy for the structural change at the redox site. The midpoint potential of cytochrome b 562 was shifted negatively by ∼135 mV by replacing Met-7 with Ala or Gly. Similar dissociation kinetics of cyanide for the mutated molecules as compared to native myoglobin was obtained.