Characterization of the precursor form of type VI collagen.
Well characterized monospecific antisera against pepsin-extracted bovine type VI collagen were used to identify and characterize the intact form of type VI collagen. In immunoblotting experiments the antisera reacted with the pepsin-resistant fragments of the alpha 1(VI) and alpha 3(VI) chains, but not with the fragment of the alpha 2(VI) chain. Extracts obtained from uterus and aorta with 6 M guanidine HCl contained two immunoreactive polypeptides of Mr = 190,000 and 180,000 based on globular protein standards. Cleavage of extracts with pepsin generated the previously characterized pepsin-resistant fragments of alpha 1(VI) and alpha 3(VI), indicating that the higher molecular weight polypeptides represent the intact parent chains, alpha 1(VI) and alpha 3(VI). Digestion of extracts with bacterial collagenase released an Mr = 100,000 noncollagenous fragment from the alpha 1(VI) chain. Thus, intact type VI collagen in tissues contains a relatively short triple helical domain and at least one very large globular domain which is sensitive to pepsin but resistant to collagenase digestion. Immunoblotting revealed a polypeptide of Mr = 240,000, which we suggest represents the pro-alpha 1(VI) chain, in the culture medium of bovine fibroblasts. Bands intermediate in molecular weight between 240,000 and 190,000 were identified in cell layers. These findings establish type VI collagen as a protein with very large nontriple helical domains, a property that undoubtedly plays an important role in its function.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001011:Aorta; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis; D005260:Female; D019791:Guanidine; D006146:Guanidines; D006801:Humans; D007106:Immune Sera; D046911:Macromolecular Substances; D003012:Microbial Collagenase; D008970:Molecular Weight; D010434:Pepsin A; D010446:Peptide Fragments / Q000032:analysis; D011347:Procollagen / Q000302:isolation & purification; D014599:Uterus
D000596:Amino Acids; D000939:Epitopes; D006146:Guanidines; D007106:Immune Sera; D046911:Macromolecular Substances; D010446:Peptide Fragments; D011347:Procollagen; D010434:Pepsin A; D003012:Microbial Collagenase; D019791:Guanidine