In order to study the effect of high-pressure (HP) treatment and two different methods of brine addition (important for lysosomal membrane destabilisation) on lysosomal enzymes activity and protein degradation, pork semitendinosus muscle was brine enhanced by injection or tumbling, and HP treated at 600 MPa following storage at 2 °C for up to 8 weeks. In this report a novel protocol for SDS gelatin zymography was established, and an increase of cathepsin B and L activity after HP treatment was shown followed by a decrease during storage. No calpain activity was detected following HP treatment. HP treatment was shown to induce a decrease in protein solubility in both myofibrillar and sarcoplasmic fractions. LC–MS analysis of these fractions showed changes in the peptide pattern during storage. Western blot analysis showed that troponin-T was indeed degraded during storage after HP treatment. The results therefore suggest that HP treatment induced an increase in cathepsin activity, which subsequently affected the myofibrillar protein degradation pattern in pork meat.