VISIBLE ABSORPTION AND ELECTRON SPIN RESONANCE SPECTRA OF ISOLATED CHAINS OF HUMAN HEMOGLOBIN . DISCUSSION OF CHAIN-MEDIATED HEME-HEME INTERACTION

The optical absorption and electron spin resonance spectra of the isolated human hemoglobin chains in the ferri state have been studied. The spectra of the isolated α+ chains are very similar to those of ferrihemoglobin, but those of the β+ chains are significantly different. The atypical spectra of the β+ chains are shown to arise from an altered spin equilibrium of the heme iron. At 0°, the aquo derivative of β+ is about 35% low spin; at liquid nitrogen temperature, the same compound is seen from electron spin resonance spectra to be 67 % low spin compared with only about 5 % for a+ and Hb+. The spin equilibrium of the β+ chains is thus greatly modified on going over to native Hb+(α2+β2+) through combination with the a+ chains. This shift is interpreted as arising from a change in the interaction between iron and histidine, resulting in a slight lengthening of the Fe-N bond. On the other hand, a similar shift in the spin equilibrium of the β+ chain is shown to occur on modifying the free thiol group next to the hemebinding histidine. Conformational strains on the peptide chain close to the heme can thus alter the spin equilibrium of the heme iron. In the reduced deoxygenated form, the β chains have a Soret band maximum at a wavelength shorter than that for a chain or ferrohemoglobin. Association with the a chain thus results in a shift of the maximum to a longer wavelength. A similar shift is observed on modifying the thiol next to the heme in the β chain. The change to longer wavelengths is in both cases associated with diminished oxygen affinity. The possibility that the Soret band shifts result from altered spin equilibrium of Fe(II) is considered. The results are discussed in the context of heme-heme interactions in native hemoglobin. © 1969, American Chemical Society. All rights reserved.