Analysis of oxygen binding to Panulirus japonicus hemocyanin. The effect of divalent cations on the allosteric transition.

The effects of H+ and divalent cations on the O2 equilibrium of hexameric hemocyanin from a spiny lobster, Panulirus japonicus, were examined. The hemocyanin showed the normal Bohr effect. When divalent cations were removed by EDTA treatment, the protein showed a fivefold increase in the O2 affinity and a considerable decrease in the cooperativity. Several cooperativity models were tested for the conformity with the observed O2-binding isotherms by the least-square curve fitting. Among the models examined, the three-state concerted model was found to be most consistent with the results. It was postulated that in the absence of divalent cations deoxyhemocyanin is mainly in the intermediate-affinity state. The arthropod hemocyanins were found to be classifiable into two groups according to their functional responses to the divalent cations. It was suggested that the cations act differently on the allosteric transitions of the two groups of hemocyanins.