Identification of a constituent of the junctional feet linking terminal cisternae to transverse tubules in skeletal muscle.
This study describes the biochemical composition of junctional feet in skeletal muscle utilizing a fraction of isolated triad junctions. [3H]Ouabain entrapment was employed as a specific marker for T-tubules. The integrity of the triad junction was assayed by the isopycnic density of [3H]ouabain activity (24-30% sucrose for free T-tubules, 38-42% sucrose for intact triads). Trypsin, chymotrypsin, and pronase all caused separation of T-tubules from terminal cisternae, indicating that the junction is composed as least in part of protein. Trypsin and chymotrypsin hydrolyzed four proteins: the Ca2+ pump, a doublet 325,000, 300,000, and an 80,000 Mr protein. T-tubules which had been labeled covalently with 125I were joined to unlabeled terminal cisternae by treatment with K cacodylate. The reformed triads were separated from free T-tubules and then severed by passage through a French press. When terminal cisternae were separated from T-tubules, some 125I label was transferred from the labeled T-tubules to the unlabeled terminal cisternae. Gel electrophoresis showed that, although T-tubules were originally labeled in a large number of different proteins, only a single protein doublet was significantly labeled in the originally unlabeled terminal cisternae. This protein pair had molecular weights of 325,000 and 300,000 daltons. Transfer of label did not occur to a substantial degree without K cacodylate treatment. We propose that the transfer of 125I label from T-tubules to terminal cisternae during reformation and breakage of the triad junction is a property of the protein which spans the gap between T-tubules and terminal cisternae.
D000818:Animals; D002101:Cacodylic Acid / Q000494:pharmacology; D064211:Connectin; D007425:Intracellular Membranes / Q000378:metabolism; D008051:Lipid Bilayers / Q000032:analysis; D008565:Membrane Proteins / Q000032:analysis; D009124:Muscle Proteins / Q000032:analysis; D009132:Muscles / Q000032:analysis / Q000378:metabolism / Q000648:ultrastructure*; D009940:Organoids / Q000032:analysis* / Q000378:metabolism; D010042:Ouabain / Q000378:metabolism; D010447:Peptide Hydrolases / Q000494:pharmacology; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000378:metabolism
D064211:Connectin; D008051:Lipid Bilayers; D008565:Membrane Proteins; D009124:Muscle Proteins; D010042:Ouabain; D002101:Cacodylic Acid; D010447:Peptide Hydrolases