A Magnetic Resonance Study of Sperm Whale Ferrimyoglobin and Its Complex with 1 Cupric Ion

Abstract

Abstract 1. Electron paramagnetic resonance (EPR) studies at 77° K of sperm whale ferrimyoglobin as a function of pH showed a transition from a predominantly high spin form to a predominantly low spin form as the pH increased, according to a pK value of about 9.2. This transition appears to coincide with the well known hemic acid dissociation observed in optical spectroscopy. The intensities of the spectra of these two forms are interpreted as accounting for the greater part of the intensities predicted for the iron(III) present. 2. A component of the signal from the more alkaline of these forms, which is tentatively labeled as high spin, is distinctly different from the high spin signal observed at neutral pH. The temperature dependence of the signals could be explained by a thermally balanced equilibrium between high spin and low spin forms, as suggested earlier for horse myoglobin. 3. Near pH 12, in a region in which the protein is known to undergo structural alteration, new high spin and low spin forms appear. These changes, like those described above, are reversed by lowering the pH. 4. The signals from 1 eq of copper(II) bound to the ferrimyoglobin were studied as a function of pH. Two forms of EPR signal were observed; they were interconverted with a pK of approximately 7.9. No intermediate forms were observed. Superhyperfine structure could be detected. The form at high pH could be recognized as highly complexed, probably with 4 nitrogen atoms involved in the coordination. The form at lower pH could not be correlated with model compounds because of the lack of entirely appropriate model studies, but it did not seem to be at variance with the conclusions of Banaszak, Kendrew, and Watson on the modes of copper(II) binding in the crystalline state. 5. Observations on complexes at pH 11 of the pentapeptide, l-valyl-l-leucyl-l-seryl-l-glutamylglycine, the amino-terminal segment of the protein, are similar to previous results with the copper(II) complexes of triglycylglycine. They show general similarities to the properties of the protein complex, but probably match it somewhat less well than does the complex with acetylglycylglycyl-l-histidylglycine. 6. In the equimolar copper(II)-ferrimyoglobin complexes there was little evidence of copper-iron interaction.