Phosphorylation inhibits the activity of μ-calpain at different incubation temperatures and Ca2+ concentrations in vitro

- The effect of phosphorylation on the activity of μ-calpain was detected in vitro.
- Incubation at 25 °C could diminish the influence of phosphorylation on μ-calpain.
- Increased Ca2+ concentration shrinks the effects caused by phosphorylation.
- Phosphorylation may play a negative role in regulating μ-calpain activity.
- The activity of m-calpain was also influenced by phosphorylation.

This study aimed to investigate the effects of phosphorylation on the activity of μ-calpain and its sensitivity to temperature and Ca2+. For temperature sensitivity analysis, sarcoplasmic protein was treated with alkaline phosphatase (AP) and phosphatase inhibitor (PI) at 4, 25 and 37 °C. The results showed that the degradation degree of μ-calpain in the AP group was significantly higher after incubation for 12 h. For calcium sensitivity analysis, samples treated with AP and PI were incubated at 0.01, 0.05, 0.1 and 1 mM Ca2+. The results showed that the degradation rate of μ-calpain was maximum in the AP group and minimum in the PI group at 0.01, 0.05 and 0.1 mM Ca2+. The differences between the three groups reduced as concentration increased. These data demonstrate that phosphorylation plays a negative role in regulating μ-calpain activity. This study clarifies the regulatory mechanism of μ-calpain activation in vitro and/or in postmortem muscle.