Cytoplasmic surface and intramembrane components of rat heart gap junctional proteins.
Gap junctions were purified from rat hearts in the presence of absence of proteolysis inhibitors and examined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and electron microscopy of thin sections. In absence of proteolysis inhibitors or in presence of ethylenediaminetetraacetic acid or leupeptin, gap junctions contained a single major protein band at relative molecular weight (Mr) 29,500 and minor bands at Mr 44,000-47,000, 17,750, and 16,500 and showed smooth cytoplasmic surfaces in electron micrographs. SDS-PAGE of junctions prepared with phenylmethylsulfonylfluoride (PMSF) showed markedly decreased intensity of the Mr 29,500 band and increased intensity of bands at Mr 44,000, 45,500, and 47,000; electron microscopy of these gap junctions showed presence of a fuzzy layer on their cytoplasmic surfaces. Urea (8 M) could not remove this fuzzy layer. In electron micrographs of rat ventricular myocytes, cytoplasmic surfaces of gap junctions were fuzzy. We conclude that rat heart gap junction protein consists of an intramembrane component (Mr 29,500) that extends into the "gap" and a cytoplasmic surface component (Mr 14,500-17,500) that corresponds to the fuzzy layer and is hydrolyzable by a serine protease.
D000818:Animals; D002458:Cell Fractionation / Q000379:methods; D002462:Cell Membrane / Q000648:ultrastructure; D004285:Dogs; D004492:Edetic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D007365:Intercellular Junctions / Q000648:ultrastructure*; D008854:Microscopy, Electron; D009206:Myocardium / Q000648:ultrastructure*; D010664:Phenylmethylsulfonyl Fluoride; D011506:Proteins / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains; D013045:Species Specificity
D011506:Proteins; D010664:Phenylmethylsulfonyl Fluoride; D004492:Edetic Acid