Enzymatic Reduction of Spin-labeled Ferrihemoglobin

• Published in: The Journal of biological chemistry Vol. 247; no. 11; pp. 3693 - 3694
• Main Authors: Asakura, T, Tamura, M, Shin, M
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 10.06.1972
• Subjects: Apoproteins; Cyclic N-Oxides; Electron Spin Resonance Spectroscopy; Ferredoxins; Hemoglobins; Humans; Iron; Methemoglobin; NAD; Oxidation-Reduction; Oxidoreductases; Oxygen; Porphyrins; Propionates; Protein Binding; Protein Conformation; Pyrrolidines; Spectrophotometry
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)45195-8

The spin-labeled ferrihemoglobin, in which a spin label is attached to one of the propionic acid groups at positions 6 or 7 of the porphyrin ring, was enzymatically reduced to the ferrous oxy form without destroying the spin label. The enzyme system contains ferredoxin, ferredoxin-NADP reductase, and a NADPH generating system. The absorption spectra of the spin-labeled hemoglobins are identical with those of native hemoglobins. The electron paramagnetic resonance spectrum of the spin-labeled oxyhemoglobin is distinctly different from that of the acid methemoglobin indicating different conformations for the protein in the vicinity of the label in these two hemoglobins. The resonance amplitude of the oxyhemoglobin is approximately twice that of the ferrihemoglobin. The change is attributed to the spin state change of the heme-iron from the high spin ferric to the low spin ferrous form. Because the oxygen binding properties of the spin-labeled hemoglobins are identical with those of native hemoglobin, this hemoglobin will be used for probing the molecular mechanism of hemoglobin functions in solution.