Human type VI collagen: Isolation and characterization of α1 and α2 chains by two-dimensional preparative gel electrophoresis

Abstract

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 α1(VI) or pepsin-derived α2(VI) chains but not with α3(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 α1 and α2 chains of type VI collagen. The globular domains of α1(VI) and α2(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 α chains may facilitate the formation of microfibrillar network.