Biosynthesis of proteoglycans in cartilage slices. Fractionation by gel chromatography and equilibrium density-gradient centrifugation.
The kinetics of incorporation of [(35)S]sulphate into slices of pig laryngeal cartilage in vitro was linear with time up to 6h. The specific radioactivities of the extracted proteoglycans (containing about 80% of the uronic acid of the cartilage) and the glycosaminoglycans remaining in the tissue after extraction were measured after various times of continuous and ;pulse-chase' radioactivity incorporation. Radioactivity was present in the isolated chondroitin sulphate after 2 min, but there was a 35min delay in its appearance in the extractable proteoglycan fraction. Fractionation of the proteoglycans by gel chromatography showed that the smallest molecules had the highest specific radioactivity, but ;pulse-chase' experiments over 5h did not demonstrate any precursor-product relationships between fractions of different size. Equilibrium density-gradient centrifugation in 4m-guanidine hydrochloride showed that among the proteoglycan fractions the specific radioactivity increased as the chondroitin sulphate content decreased, but with preparations from ;pulse-chase' experiments there was again no evidence for precursor-product relationships between the different fractions. Differences in radioactive incorporation would seem to reflect metabolic heterogeneity within the proteoglycans extracted from cartilage. This may be due either to a partial separation of different types of proteoglycans or to differences in the rates of degradation of the molecules of different size and composition as a result of the nature and specificity of the normal degrading enzymes. The results suggest that molecules of all sizes were formed at the same time.
D000818:Animals; D002499:Centrifugation, Density Gradient; D002845:Chromatography; D006023:Glycoproteins / Q000096:biosynthesis*; D006025:Glycosaminoglycans / Q000096:biosynthesis*; D006146:Guanidines; D066298:In Vitro Techniques; D007700:Kinetics; D007817:Laryngeal Cartilages / Q000378:metabolism*; D008722:Methods; D013431:Sulfates / Q000378:metabolism; D013460:Sulfur Isotopes; D013552:Swine; D014574:Uronic Acids / Q000032:analysis
D006023:Glycoproteins; D006025:Glycosaminoglycans; D006146:Guanidines; D013431:Sulfates; D013460:Sulfur Isotopes; D014574:Uronic Acids