Isolation of the N-terminal globular protein domains from cartilage proteoglycans. Identification of G2 domain and its lack of interaction with hyaluronate and link protein.
The N-terminal fragment (G1-G2) of cartilage proteoglycan protein core contains two globular domains, binding region (G1) and a second globular domain (G2), G1-G2 was isolated after mild trypsin digestion of purified proteoglycan aggregates followed by chromatography first on Sepharose CL-2B under associative conditions and then on a TSK-4000 column in 4 M-guanidinium chloride. It migrated as a single band (apparent Mr 150,000) on SDS/polyacrylamide-gel electrophoresis. G2 was isolated by V8-proteinase digestion of G1-G2 followed by aggregation of the G1-containing fragments with hyaluronate and chromatography on TSK-4000. It migrated as a single band on SDS/polyacrylamide-gel electrophoresis of apparent Mr 66,000 after digestion with keratanase. G2 did not interact with proteoglycan monomer, hyaluronate, link protein or other extractable cartilage matrix proteins. A polyclonal antibody raised against G2 did not cross-react with G1 or link protein. These data show that, despite a high degree of sequence similarity, G1 and G2 do not share any functional properties nor have major antigenic sites in common.
Fosang
A J
AJ
Biochemistry Division, Kennedy Institute of Rheumatology, Hammersmith, London, U.K.
Hardingham
T E
TE
D053674:Aggrecans; D000818:Animals; D011508:Chondroitin Sulfate Proteoglycans / Q000302:isolation & purification / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins / Q000302:isolation & purification* / Q000378:metabolism; D006820:Hyaluronic Acid / Q000378:metabolism*; D007817:Laryngeal Cartilages / Q000032:analysis*; D037181:Lectins, C-Type; D011506:Proteins / Q000378:metabolism*; D011509:Proteoglycans; D012697:Serine Endopeptidases; D013552:Swine; D000099249:Proteoglycan Link Protein
D053674:Aggrecans; D011508:Chondroitin Sulfate Proteoglycans; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D006820:Hyaluronic Acid; D037181:Lectins, C-Type; D011506:Proteins; D011509:Proteoglycans; D012697:Serine Endopeptidases; D000099249:Proteoglycan Link Protein; C028344:glutamyl endopeptidase