Loss of Ca²⁺-dependent Regulation in Glycerinated Skeletal Muscle Contraction

Abstract

Glycerinated muscle fiber from rabbit psoas muscle often lost Ca²⁺-dependent regulation of its contraction with long-term extraction in a 50% glycerol solution containing 5mM EGTA at-20°C, designated as Ca²⁺-insensitive muscle fiber (CaIS-fiber). About 30 or 40% of glycerinated muscle fibers were CaIS-fibers after 1 to 3 months in the glycerol solution. We investigated the cause of the loss of Ca²⁺-sensitivity of the glycerinated muscle fiber by tension mechanogram and SDS polyacrylamide gel electrophoresis.
This natural CaIS-fiber showed a new band of 30K daltons peptide on SDS gels. On the other hand, Ca²⁺-sensitive fiber (CaS-fiber) changed to CaIS-fiber by trypsin digestion for 40sec. The tryptic CaIS-fiber had no troponin C and 30K daltons peptide bands in the electrophoretograms. Incubating with 2mM CaCl₂ for 40hr at 25°C, CaS-fiber changed easily to CaIS-fiber which had 30K daltons peptide and faint troponin T and I bands, as in natural CaIS-fiber. All CaIS-fibers could recover their Ca²⁺-dependent regulation by incubating with native tropomyosin from rabbit skeletal muscle for 2 days at 4°C.
These results indicate that the loss of Ca²⁺-dependent regulation of glycerinated muscle fiber is due to degradation of regulatory protein system by endogenous Ca²⁺-activated proteolytic enzymes.