Biochemical and physiochemical characterization of pepsin-solubilized type-II collagen from bovine articular cartilage

Abstract

Solubilization of collagen from bovine articular cartilage with pepsin requires the preliminary extraction of proteoglycans from the ground substance. Biochemical and physicochemical properties of this pepsin-solubilized collagen are independent of the pretreatment (extraction with 1.5 M-CaCl2, 5 M-guanidinium chloride or 0.2 M-NaOH) and of the age range (2-4 yr old and 2 mo. old animals). Characterization of the denatured components, of the CNBr peptides and of the amino acid and cross-link composition shows that the collagen of the hyaline cartilage is all type II. Electrical birefringence measurements showed the presence of tropocollagen molecules (length 280 nm) and molecules whose length is slightly less than twice that of the tropocollagen molecules. This latter molecule may be a dimer composed of 2 monomers linked by intermolecular head-to-tail bonds and whose theoretical length (530 nm), according to the quarter-stagger theory, is in good agreement with measured values (510-530 nm). It was verified that the .beta.-components of this collagen are formed of 2 .alpha.-chains linked by the stable intermolecular bond, dehydrodihydroxylysinonorleucine. These dimeric molecules are absent from solutions of skin collagen whose .beta.-components possess only aldol-type intramolecular cross-links. Although reconstituted fibers from solutions of skin and cartilage collagen are similar, the segment-long spacing crystallites formed with pepsin-solubilized cartilage collagen present a symmetrical and dimeric form corresponding to the lateral aggregation of 2 monomers with an overlap (90 nm) of the C-terminal ends.