Isolation and characterization of calcium binding glycoproteins of cardiac sarcolemmal vesicles.
Two major Ca2(+)-binding glycoproteins Mr 120,000 and 100,000 were isolated from 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid -solubilized bovine heart sarcolemma membrane. Peroxidase-conjugated concanavalin A and wheat germ agglutinin lectins bind strongly to the isolated 120- and 100-kDa glycoproteins. Treatment with endoglycosidase F resulted in conversion of the 120-kDa glycoprotein to a form migrating at about 97 kDa. Treatment of the 100-kDa band with endoglycosidase F produced form of about 80 kDa. Endoglycosidase H digestion removes only 5% of the mass of both glycoproteins. the carbohydrate structure of both glycoproteins, is therefore, predicted to be at least 75% complex structure and 25% high mannose or hybrid structure. The 120- and 100-kDa glycoproteins are the major Ca2(+)-binding proteins in the sarcolemma membranes. Intact and endoglycosidase-treated glycoproteins bind 45Ca2+ as analyzed by a 45Ca2+ overlay technique. Using polyclonal antibodies, the 120- and 100-kDa glycoproteins were identified in muscle plasma membranes (ventricles, atria, and uterus smooth muscle). They were, however, not present in non-muscle tissues such as pancreas, liver, and kidney. The 120- and 100-kDa glycoproteins appear to be homologous molecules as judged by their similar V8 protease peptide maps, cross-reactivity with polyclonal antibody, and other physicochemical properties.
Michalak
M
M
Cardiovascular Disease Research Group, University of Alberta, Edmonton, Canada.
Fliegel
L
L
Wlasichuk
K
K
D000118:Acetylglucosaminidase / Q000378:metabolism; D000818:Animals; D002132:Calcium Radioisotopes / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002417:Cattle; D002462:Cell Membrane / Q000032:analysis; D002846:Chromatography, Affinity; D003208:Concanavalin A / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000302:isolation & purification* / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000378:metabolism; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D008970:Molecular Weight; D009130:Muscle, Smooth / Q000032:analysis; D009206:Myocardium / Q000032:analysis*; D010449:Peptide Mapping; D012508:Sarcolemma / Q000032:analysis*; D012697:Serine Endopeptidases / Q000378:metabolism; D014018:Tissue Distribution; D014909:Wheat Germ Agglutinins / Q000378:metabolism
D002132:Calcium Radioisotopes; D002135:Calcium-Binding Proteins; D006023:Glycoproteins; D014909:Wheat Germ Agglutinins; D003208:Concanavalin A; D006026:Glycoside Hydrolases; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase