Aplysia oxymyoglobin with an unusual stability property: involvement of two kinds of car☐yl groups

Abstract

Unlike mammalian oxymyoglobins, Aplysia MbO 2 is extremely susceptible to autoxidation, and its pH dependence is also unusual. Kinetic formulation has revealed that two kinds of dissociable group with p K 1 = 4.3 and, p K 2 = 6.1, respectively, at 25°C are involved in the stability property of Aplysia MbO 2. In order to characterize thermodynamically these dissociation processes involved, the effect of temperature on K 1 and K 2 was studied by analyzing the pH dependence for the autoxidation rate of Aplysia MbO 2 in 0.1 M buffer over the pH range of 4–11, and at 15, 25 and 35°C. The resulting thermodynamic parameters for each group were both those to be expected for the ionization of a car☐yl group; the ΔH° value being numerically much less than 1 kcal·mol −1, or zero in practice, but being associated with a large negative values of ΔS° of the order of −20 cal·mol −1·K −1. Taking into account the fact that Aplysia myoglobin contains only a single histidine residue corresponding to the heme-binding proximal one, we can unequivocally conclude that the two kinds of the dissociable group involved in the unusual stability of Aplysia MbO 2 must both be car☐yl groups, the protonation of these groups being responsible for an increase in its autoxidation rate in the acidic pH range.