Differential sensitivity of chicken MM-creatine kinase to trypsin and proteinase-K

Under several conditions of SDS-PAGE, the chicken MM-creatine kinase (MM-CK) monomer migrated as a approximately 50,000 dalton polypeptide, approx 25% larger than usually reported. Characterization by sedimentation equilibrium indicated that the anomalous molecular weight was an artifact of electrophoresis. Digestion with trypsin caused only moderate reductions in CK activity, despite extensive degradation of the denatured enzyme revealed by SDS-PAGE. Characterization of trypsinized MM-CK under non-denaturing conditions of electrophoresis and HPLC revealed no fragmentation of the native enzyme, suggesting that MM-CK quaternary structure was maintained despite extensive tryptic nicking. In contrast, much lower concentrations of proteinase-K generated only a single fragment in SDS-PAGE while causing a nearly total loss of enzyme activity.