Postmortem changes in myofibrillar-bound calpain 3 revealed by immunofluorescence microscopy

Area of Science:

• Muscle biology
• Proteolysis
• Meat science

Background:

• Calpains are calcium-dependent proteases involved in muscle protein turnover.
• Understanding postmortem changes in meat quality is crucial for the food industry.
• Calpain 3 (p94) plays a significant role in muscle structure and function.

Purpose of the Study:

• To develop a method for monitoring myofibrillar-bound calpain 3 changes.
• To investigate the proteolytic activity of calpains 1 and 3 during postmortem storage.
• To examine the breakdown of titin and nebulin in relation to calpain activation.

Main Methods:

• Development of an immunofluorescence microscopy technique for calpain 3.
• Western blot analysis of calpain 3, calpain 1, titin, and nebulin.
• Examination of ovine longissimus thoracis et lumborum (LTL) myofibrils stored for 0-3 days postmortem.

Main Results:

• Calpain 3 levels significantly decreased postmortem, becoming undetectable by day 3.
• Calpain 1 was converted from an 80 kDa intact form to a 76 kDa fragment.
• Calpain 3 localized to Z- and M-lines, with fluorescence intensity declining during storage.
• Titin and nebulin breakdown paralleled the decrease in calpain 3 fluorescence.

Conclusions:

• Calpains 1 and 3 are activated during postmortem storage of ovine meat.
• Activated calpains contribute to the proteolytic degradation of myofibrillar proteins, impacting meat quality.
• Immunofluorescence microscopy is effective for tracking calpain 3 activity in postmortem muscle.