Type VI collagen: high yields of a molecule with multiple forms of alpha 3 chain from avian and human tissues.
A differential extraction procedure followed by molecular sieve column chromatography for the isolation of large quantities of the tissue form of type VI collagen is described. Recovery of the protein was more than 60% from both chick gizzard and human placenta. On reduced NaDodSO4-gels chick type VI collagen migrated as two major bands at Mr = 140,000 and 150,000 that were present in a 1:1 ratio and five less intense bands between Mr = 230,000 and 180,000. By immunoblotting with a polyclonal antibody against the pepsinized form of chick type VI collagen, all these bands were stained. Furthermore, the amino acid composition of the five higher Mr polypeptides indicated that they all contained hydroxyproline and hydroxylysine. In the chick type VI collagen molecule the five bands of higher Mr belong to the alpha 3 chain since they were recognized by monoclonal antibodies specific for the chick Mr = 260,000 alpha 3 chain. On examination of antigenic activity by solid-phase radioimmunobinding, densitometry of stained NaDodSO4 polyacrylamide gels, and protein content type VI was found to be an abundant collagen since it accounted for up to 0.1% of the tissue wet weight. The yields per tissue wet weight and the migration pattern of human type VI collagen polypeptides were similar to those of the chick. Agarose/polyacrylamide composite gels indicated that the molecular size of the tissue form of type VI collagen molecules under non-reduced conditions corresponded to a basic type of tetrameric molecule.
Divisione di Oncologia Sperimentale 2, Centro di Riferimento Oncologico, Aviano, Italy.
A
A
Colombatti
K
K
Ainger
F
F
Colizzi
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D015153:Blotting, Western; D002642:Chick Embryo; D002850:Chromatography, Gel; D003094:Collagen / Q000276:immunology / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D005109:Extracellular Matrix / Q000032:analysis*; D005895:Gizzard, Avian / Q000032:analysis; D006801:Humans; D008970:Molecular Weight; D010446:Peptide Fragments / Q000276:immunology / Q000302:isolation & purification*; D010920:Placenta / Q000032:analysis
D000596:Amino Acids; D010446:Peptide Fragments; D003094:Collagen