Effect of protein S-nitrosylation on autolysis and catalytic ability of μ-calpain

The effect of S-nitrosylation on the autolysis and catalytic ability of l-calpain in vitro in the presence of 50 mu M Ca2 + was investigated. mu-Calpain was incubated with different concentrations of nitric oxide donor S-nitrosoglutathione (GSNO) and subsequently reacted with purified myofibrils. Results showed that the amount of 80 kDa mu-calpain subunit significantly decreased as GSNO increased from 0 to 300 mu M, but increases of GSNO to 300, 500 and 1000 mu M did not result in further inhibition. The catalytic ability of nitrosylated mu-calpain to degrade titin, nebulin, troponin-T and desmin was significantly reduced when the GSNO concentration was higher than 300 mu M. The cysteine residues of l-calpain at positions 49, 351, 384, and 592 in the catalytic subunit and at 142 in small subunit were S-nitrosylated, which could be responsible for decreased mu-calpain activity. Thus, S-nitrosylation can negatively regulate the activation of mu-calpain resulting in decreased proteolytic ability on myofibrils. (C) 2016 Elsevier Ltd. All rights reserved.