Eastern grey kangaroo muscle creatine kinase.

Creatine kinase has been purified to homogeneity from skeletal muscle of the eastern grey kangaroo, Macropus giganteus. The procedure included ethanol fractionation followed by chromatography on DEAE-cellulose. The enzyme had a molecular weight of approximately 86 000 with two subunits of 43 500. Two sulfhydryl groups were determined for the intact molecule and a further four on unfolding. Under standardized conditions, the metal ion specificity was determined as MgADP- greater than MnADP- greater than CoADP-, CaADP-; and the substrate specificity as MgADP- greater than MgdADP- greater than MgGDP- greater than MgXDP. Initial velocity and product-inhibition studies of the reverse reaction were consistent with a rapid random equilibrium reaction as observed for the rabbit muscle enzyme. However, initial-velocity studies in the forward reaction were consistent with a rapid equilibrium-ordered mechanism with MgATP2- binding before creatine. Preliminary studies on the binding of manganese nucleotides to the enzyme have been carried out using pulsed nuclear magnetic resonance to measure relaxation times of water protons.