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Updated: Feb 3, 2026

Measuring Calpain Activity in Fixed and Living Cells by Flow Cytometry
Published on: July 8, 2010
Calpastatin inhibits the activity of phosphorylated μ-calpain in vitro
Manting Du1, Xin Li2, Zheng Li2
1Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Key Laboratory of Agro-Product Processing, Ministry of Agriculture, Beijing 100193, PR China; College of Food and Biological Engineering, Zhengzhou University of Light Industry, Zhengzhou, Henan 450000, PR China.
Abstract:
The objective of this study was to investigate the effect of phosphorylation on the sensitivity of μ-calpain to the inhibition induced by calpastatin. Purified μ-calpain was incubated with alkaline phosphatase (AP) or protein kinase A (PKA) to modulate the phosphorylation level of μ-calpain. Accurately 25, 50, 100 and 150 units of AP/PKA-treated μ-calpain were mixed with the same amounts of heat stable proteins and incubated at 4 °C. In the calpastatin-free system, AP and PKA-treated μ-calpain had higher proteolytic activity compared to the control. Intact AP-treated μ-calpain degraded fastest in the 50, 100 and 150 unit μ-calpain incubation systems. However, the degradation rate of μ-calpain in control and PKA group was non-significant in 100 and 150 unit μ-calpain systems. Our results demonstrated that, compared to dephosphorylated and control μ-calpain, calpastatin presents greater inhibition to PKA phosphorylated μ-calpain. This study increases understanding of the mechanism of μ-calpain activity regulated by phosphorylation.
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