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

Abstract

Solubilization of collagen from bovine articular with pepsin requires the preliminary extraction of proteoglycans from the ground substance. Biochemical and physiochemical properties of this pepsin-solubilized collagen are independent of the pretreatment (extraction with 1.5M-CaCl2, 5M-guanidinium chloride or 0.2M-NaOH) and of the age range (2-4-year-old and 2-month-old animals). Characterization of the de-natured 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 280nm) and molecules whose length is slightly less than twice that of the tropocollagen molecules. This latter molecule may be a dimer composed of two monomers linked by intermolecular head-to-tail bonds and whose theoretical length (530nm), according to the quarter-stagger theory, is in good agreement with our measured values (510-530nm). We have verified that the beta-components of this collagen are formed of two 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 fibres 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 two monomers with an overlap (90nm) of the C-terminal ends.

Mechanisms of cartilage growth: Modulation of balance between proteoglycan and collagen in vitro using chondroitinase ABC

“…DNA was converted to cell number by using a conversion constant of 7.7 pg of DNA per cell (36). Hydroxyproline was converted to collagen by assuming a mass ratio of collagen: hydroxyproline equal to 7.25:1 (17,37). Portions of spent medium were analyzed for GAG (33).…”

Articular cartilage tensile integrity: Modulation by matrix depletion is maturation-dependent

“…In experiment 2, HA content was determined for individual samples. Hydroxyproline content was converted to COL content by assuming a mass ratio of collagen to hydroxyproline equal to 7.25 for bovine cartilage [58,59] and 7.1 for human cartilage [60]. The molar ratio of PYR per collagen molecule was calculated, assuming the molecular weight of collagen triple helix of 300,000.…”

Cockroach Collagen: Isolation, Biochemical and Biophysical Characterization

“…The values obtained are presented in Table 1 and compared with the composition of collagen rat tail (type I) and bovine articular cartilage (type 11) [28]. The analysis exhibits all the characteristic features of a collagenous protein : glycine typically makes up almost one-third of all the amino acid residues, and the imino acids (proline and hydroxyproline) are in relatively high concentration (23 %).…”

“…However, the first determinations of the amino acid composition showed a contamination by non-collagenous components. Therefore, prior to enzymatic digestion, an extraction with 4 M guanidinium chloride was performed as described for vertebrate tissues [18,28]. This pretreatment extracted 3% of the total collagen, but increased the quantity of extractable pepsin collagen up to 66% of the total.…”