Purification and N-terminal amino acid sequence of a chondroitin sulphate/dermatan sulphate proteoglycan isolated from intima/media preparations of human aorta.
A proteoglycan (PG) was purified to homogeneity from intima/media preparations of human aorta specimens by the following chromatographic steps: Sepharose Q anion exchange, Sepharose CL-4B size exclusion, hydroxyapatite, MonoQ anion exchange and TSK G 4000 SW size exclusion. The purity of the preparation was established by SDS/PAGE using direct staining by silver or Dimethylmethylene Blue, as well as by Western blots of biotin-labelled samples. The electrophoretic mobility of the native PG was less than that of a 200,000-Mr standard protein. After treatment with chondroitin sulphate lyase ABC, a core protein of Mr 15,000 was revealed. The Mr of the glycosaminoglycan (GAG) peptides was less than 24,000, by comparison with a keratan sulphate peptide. The composition of the GAG chains was determined by differential digestion of the PG by chondroitin sulphate lyases AC/ABC or chondroitin sulphate lyase AC alone followed by anion-exchange chromatography of the resulting disaccharides. The GAG chains are composed of approximately one-third of dermatan sulphate and two-thirds chondroitin sulphate disaccharide units. The sequence of the 20 N-terminal amino acids is identical with the sequence previously reported for PG I isolated from human developing bone [Fisher, Termine & Young (1989) J. Biol. Chem. 264, 4571-4576]. The assignment of glycosylation sites to the serine residues in positions 5 and 10 was confirmed. The findings indicate that the chondroitin sulphate/dermatan sulphate PG is a major PG in intima/media preparations of human aorta and represents a biglycan-type PG.
Stöcker
G
G
Department of Clinical Chemistry and Pathobiochemistry, Medical Faculty, University of Technology, Aacher, Federal Republic of Germany.
Meyer
H E
HE
Wagener
C
C
Greiling
H
H
D000595:Amino Acid Sequence; D001011:Aorta / Q000737:chemistry*; D011508:Chondroitin Sulfate Proteoglycans / Q000737:chemistry / Q000302:isolation & purification*; D002845:Chromatography; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003871:Dermatan Sulfate / Q000737:chemistry / Q000302:isolation & purification*; D017886:Durapatite; D006595:Hexosamines / Q000032:analysis; D006801:Humans; D006882:Hydroxyapatites; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009131:Muscle, Smooth, Vascular / Q000737:chemistry*
D011508:Chondroitin Sulfate Proteoglycans; D006595:Hexosamines; D006882:Hydroxyapatites; C032649:dermatan sulfate proteoglycan; D003871:Dermatan Sulfate; D017886:Durapatite