Human type VI collagen: isolation and characterization of alpha 1 and alpha 2 chains by two-dimensional preparative gel electrophoresis.
Two 140 kDa collagenous glycoproteins were isolated from 5 M guanidinium chloride extracts of human uterine leiomyoma by two-dimensional preparative gel electrophoresis. The glycoproteins represented the major concanavalin A binding fraction of the extract and were also present in adult human skin. On two-dimensional gel electrophoresis the glycoproteins appeared as elongated spots, indicating variations of their isoelectric points from 5 to 6. These glycoproteins were disulfide-bonded components of high molecular mass protein and, after reduction, became sensitive to collagenase treatment that generated peptides corresponding in size to those of the noncollagenous domains of type VI collagen. Antisera raised against these purified glycoproteins reacted with either pepsin-derived alpha 1(VI) or pepsin-derived alpha 2(VI) chains but not with alpha 3(VI) chain of human type VI collagen. Reciprocally, these glycoproteins reacted with monoclonal antibodies against type VI collagen. These results indicate that the glycoproteins represent the integral alpha 1 and alpha 2 chains of type VI collagen. The globular domains of alpha 1(VI) and alpha 2(VI) chains remaining after collagenase treatment appeared on two-dimensional gel electrophoresis as elongated spots, suggesting that the noncollagenous portions determine the well known microheterogeneity of the molecule. The differences in isoelectric points between and within alpha chains may facilitate the formation of microfibrillar network.
Wegrowski
J
J
Laboratory of Biochemistry, CNRS URA 610, Faculty of Medicine, University of Reims, France.
Bellon
G
G
Randoux
A
A
D000596:Amino Acids / Q000032:analysis; D000911:Antibodies, Monoclonal; D003094:Collagen / Q000302:isolation & purification*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D005260:Female; D019791:Guanidine; D006146:Guanidines; D006801:Humans; D007889:Leiomyoma / Q000032:analysis; D046911:Macromolecular Substances; D008970:Molecular Weight; D011247:Pregnancy; D012867:Skin / Q000032:analysis; D014594:Uterine Neoplasms / Q000032:analysis
D000596:Amino Acids; D000911:Antibodies, Monoclonal; D006146:Guanidines; D046911:Macromolecular Substances; D003094:Collagen; D019791:Guanidine