Isolation and characterization of a 115,000-dalton matrix-associated glycoprotein from chick aorta.
Chick aortas were extracted sequentially with phosphate-buffered saline, 6 M guanidine HCl, and 6 M guanidine HCl containing dithioerythritol. The proteins present in the guanidine HCl + dithioerythritol extract were separated by DEAE-cellulose chromatography, and the fractions recovered were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Five major glycoprotein components with apparent Mr = 205,000, 195,000, 150,000, 135,000, and 115,000 (gp 115) were identified. gp 115 was further studied since it was the only noncollagenous protein based on amino acid analysis. The protein was purified to homogeneity by preparative electrophoresis. Its amino acid composition was characterized by a high content of glutamic acid and arginine and a relatively high content of leucine, glycine, and alanine. The concentration of gp 115 in the guanidine HCl + dithioerythritol extract was about 15-fold that in the guanidine and saline extracts. Overall, about 80% of the protein was solubilized with guanidine HCl + dithioerythritol, suggesting that most of it formed large aggregates stabilized by disulfide bonds in vivo. Immunofluorescence studies with specific antibodies showed that gp 115 formed an extracellular fibrillar network in the aorta wall. One-dimensional finger printing with Staphylococcus aureus V8 protease and immunological studies indicated that the protein was unrelated to fibronectin and laminin. The data led us to conclude that gp 115 is a novel extracellular component of chick aorta.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D000936:Antigen-Antibody Complex; D001011:Aorta / Q000032:analysis*; D002645:Chickens; D004226:Dithioerythritol; D005109:Extracellular Matrix / Q000032:analysis*; D005455:Fluorescent Antibody Technique; D006023:Glycoproteins / Q000032:analysis*; D019791:Guanidine; D006146:Guanidines; D007106:Immune Sera; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis
D000596:Amino Acids; D000936:Antigen-Antibody Complex; D006023:Glycoproteins; D006146:Guanidines; D007106:Immune Sera; D010446:Peptide Fragments; D004226:Dithioerythritol; D019791:Guanidine