Characterization of proteoglycans isolated from associative extracts of human articular cartilage.
Approx. 10% of the total proteoglycan content of normal young human articular cartilage was extracted under associative conditions with Dulbecco's PBS. Proteoglycans isolated from the extract by Q-Sepharose chromatography were separated by gel chromatography and characterized by gradient gel SDS/PAGE and immunoblotting. Three species of small proteoglycans, two main populations of aggrecan and a population of its smaller fragments were identified. The major populations of aggrecan contained chondroitin sulphate chains, all or part of the N-terminal G1 and G2 domains and, therefore, intact keratan sulphate domains. The larger population was estimated by gradient SDS/PAGE to have a molecular mass of approx. 600 kDa or greater. The second population had an apparent molecular mass of approx. 300-600 kDa. Core proteins derived from these populations of proteoglycans separated on SDS/PAGE into several clusters of bands in the range from 120 to approx. 360 kDa. The extract further contained smaller fragments which lacked chondroitin sulphate but reacted with antibodies against keratan sulphate, and against epitopes present in the G2 domain of aggrecan. The presence of the G2 domain in a broad range of populations of decreasing size indicated extensive cleavage of the aggrecan core protein within its chondroitin sulphate domain. These findings suggest that fragmentation of aggrecan probably occurs in vivo in normal articular cartilage of young individuals. Associative extracts also contained decorin, biglycan and fibromodulin. These were resolved from aggrecan by gel chromatography and identified by immunodetection.
Vilím
V
V
Institute of Rheumatology, Praha, Czech Republic.
Fosang
A J
AJ
D053674:Aggrecans; D000911:Antibodies, Monoclonal; D002358:Cartilage, Articular / Q000378:metabolism*; D005591:Chemical Fractionation; D002807:Chondroitin / Q000737:chemistry; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D016326:Extracellular Matrix Proteins; D006801:Humans; D015151:Immunoblotting; D037181:Lectins, C-Type; D008970:Molecular Weight; D011509:Proteoglycans / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism
D053674:Aggrecans; D000911:Antibodies, Monoclonal; D016326:Extracellular Matrix Proteins; D037181:Lectins, C-Type; D011509:Proteoglycans; D002807:Chondroitin