Immobilized apo-myoglobin, a new stable reagent for measuring rates of heme dissociation from hemoglobin.

Apo-myoglobin covalently linked on CNBr-activated Sepharose 4B is proposed as a new heme acceptor for investigating the heme transfer reaction from hemoproteins. Immobilized apo-myoglobin has the desirable properties of an ideal heme acceptor in that it is characterized by a high affinity for ferric heme, a high stability towards denaturation even at physiological temperatures and can be lyophilized for long-term storage. The study of heme release from myoglobin at pH 5.0 and 37 degrees C indicates that heme affinity is increased at least 10-fold relative to the soluble protein. Experiments with human hemoglobin allowed the estimation of the heme release rates from both alpha and beta chains and brought out the greater temperature sensitivity of the alpha chain heme-globin linkage.