The erythrocruorin of Eisenia fetida. I. Properties and subunit structure.

The erythrocruorin of Eisenia fetida possesses a relative molecular mass, determined by sedimentation equilibrium, of (3.82 +/- 0.05) . 10(6). According to the iron and heme contents, 0.218 +/- 0.008% and 2.34 +/- 0.02% by mass, respectively, it contains 144 hemes per molecule. The dimensions of the molecule observed by electron microscopy are 25.0 X 16.5 nm (diameter X height). SDS-polyacrylamide gel electrophoresis indicates that the erythrocruorin consists of six subunits (Mr 14,900, 15,300, 17,200, 19,700, 31,600 and 40,000). Oxygen binding studies showed that E. fetida erythrocruorin has a high oxygen affinity (P50 = 2.8 Torr at pH 7.5), exhibits a slight bohr effect and possesses a high cooperativity with the Hill coefficient h = 3.7-4.8. Treatment of the erythrocruorin either by freezing and thawing or by aging or exposure to alkaline pH converts it irreversibly into a state of lower cooperativity with h = 2.0-2.6. A model of the subunit structure of the erythrocruorin is proposed which takes into account the physiochemical and oxygen-binding properties of the erythrocruorin and the subunits obtained upon its dissociation.