Structural electrochemical study of hemoglobin by in situ circular dichroism thin layer spectroelectrochemistry.

Secondary and tertiary or quaternary structural changes in hemoglobin (HB) during an electroreduction process were studied by in situ circular dichroism (CD) spectroelectrochemistry with a long optical path thin-layer cell. By means of singular value decomposition least-squares analysis, CD spectra in the far-UV region give two similar alpha components with different CD intensity, indicating slight denaturation in the secondary structures due to the electric field effect. CD spectra in the Soret band show a R-->T transition of two quaternary structural components induced by electroreduction of the heme, which changes the redox states of the center ion from Fe3+ to Fe2+ and the co-ordination number from 6 to 5. The double logarithmic analysis shows that electroreduction of hemoglobin follows a chemical reaction with R-->T transition. Some parameters in the electrochemical process were obtained: formal potential, E0'=-0.167 V; electrochemical kinetic overpotential, deltaE0=-0.32 V; standard electrochemical reaction rate constant, k0=1.79 x 10(-5) cm s(-1); product of electron transfer coefficient and electron number, alphan=0.14; and the equilibrium constant of R-->T transition, Kc=9.0.