Biochemical properties of purified transverse tubules isolated from skeletal muscle triads.
Transverse tubules (t-tubules) were prepared from muscle by dissociation of intact triads during centrifugation in ion-free sucrose gradients. They were further purified by the removal of contaminating sarcoplasmic reticulum after loading with calcium phosphate. Purification was accompanied by enrichment in markers specific for t-tubules, e.g., nitrendipine binding sites. According to gel electrophoresis the purified t-tubules contained three major protein bands of 104, 70, and 30 kDa. When solubilized with detergents there was a two- to threefold increase in Mg2+-ATPase activity, and a corresponding increase in the 30-kDa protein band. The 104-kDa protein was shown to be a (Na+ + K+)-ATPase because of its phosphorylation by [gamma-32P]ATP in the presence of sodium ions. The orientation of the t-tubule membrane was predominantly inside-out.
Horgan
D J
DJ
CSIRO Division of Food Research, Queensland, Australia.
Kuypers
R
R
D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000302:isolation & purification; D000252:Calcium-Transporting ATPases / Q000302:isolation & purification; D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D007425:Intracellular Membranes / Q000201:enzymology; D008870:Microtubules / Q000032:analysis* / Q000201:enzymology; D008970:Molecular Weight; D009124:Muscle Proteins / Q000302:isolation & purification; D009132:Muscles / Q000032:analysis* / Q000166:cytology / Q000201:enzymology; D010744:Phosphoric Monoester Hydrolases / Q000302:isolation & purification; D010766:Phosphorylation; D012519:Sarcoplasmic Reticulum / Q000201:enzymology
D009124:Muscle Proteins; D010744:Phosphoric Monoester Hydrolases; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases