Interaction of dimeric horse cytochrome c with cyanide ion.

We have previously shown that methionine-heme iron coordination is perturbed in domain-swapped dimeric horse cytochrome c. To gain insight into the effect of methionine dissociation in dimeric cytochrome c, we investigated its interaction with cyanide ion. We found that the Soret and Q bands of oxidized dimeric cytochrome c at 406.5 and 529 nm redshift to 413 and 536 nm, respectively, on addition of 1 mM cyanide ion. The binding constant of dimeric cytochrome c and cyanide ion was obtained as 2.5 × 10(4) M(-1). The Fe-CN and C-N stretching (ν (Fe-CN) and ν (CN)) resonance Raman bands of CN(-)-bound dimeric cytochrome c were observed at 443 and 2,126 cm(-1), respectively. The ν (Fe-CN) frequency of dimeric cytochrome c was relatively low compared with that of other CN(-)-bound heme proteins, and a relatively strong coupling between the Fe-C-N bending and porphyrin vibrations was observed in the 350-450-cm(-1) region. The low ν (Fe-CN) frequency suggests weaker binding of the cyanide ion to dimeric cytochrome c compared with other heme proteins possessing a distal heme cavity. Although the secondary structure of dimeric cytochrome c did not change on addition of cyanide ion according to circular dichroism measurements, the dimer dissociation rate at 45 °C increased from (8.9 ± 0.7) × 10(-6) to (3.8 ± 0.2) × 10(-5) s(-1), with a decrease of about 2 °C in its dissociation temperature obtained with differential scanning calorimetry. The results show that diatomic ligands may bind to the heme iron of dimeric cytochrome c and affect its stability.