A rat osteogenic cell line (UMR 106-01) synthesizes a highly sulfated form of bone sialoprotein

• Published in: The Journal of biological chemistry Vol. 265; no. 9; pp. 5285 - 5291
• Main Authors: MIDURA, R. J, MCQUILLAN, D. J, BENHAM, K. J, FISHER, L. W, HASCALL, V. C
• Format: Journal Article
• Language: English
• Published: Bethesda, MD American Society for Biochemistry and Molecular Biology 25.03.1990
• Subjects: 550201 - Biochemistry- Tracer Techniques; 550901 - Pathology- Tracer Techniques; AMINES; AMINO ACID SEQUENCE; AMINO ACIDS; Analytical, structural and metabolic biochemistry; ANIMAL CELLS; ANIMALS; BASIC BIOLOGICAL SCIENCES; BETA DECAY RADIOISOTOPES; BETA-MINUS DECAY RADIOISOTOPES; Biological and medical sciences; BIOSYNTHESIS; CARBOHYDRATES; CARBOXYLIC ACIDS; Cell Line; CHROMATOGRAPHY; Chromatography, Gel; DAYS LIVING RADIOISOTOPES; DISEASES; EVEN-ODD NUCLEI; Fundamental and applied biological sciences. Psychology; GLUCOSAMINE; Glucosamine - metabolism; GLYCOPROTEINS; HEXOSAMINES; HEXOSES; HYDROGEN COMPOUNDS; HYDROXY ACIDS; ISOTOPE APPLICATIONS; ISOTOPES; LIGHT NUCLEI; MAMMALS; MOLECULAR STRUCTURE; MONOSACCHARIDES; NEOPLASMS; NUCLEI; OLIGOSACCHARIDES; ORGANIC ACIDS; ORGANIC COMPOUNDS; osteogenesis; Osteopontin; Osteosarcoma; OSTEOSARCOMAS; OXYGEN COMPOUNDS; PRECURSOR; PROTEINS; RADIOISOTOPES; RATS; RODENTS; SACCHARIDES; SARCOMAS; SECRETION; SEPARATION PROCESSES; Sialoglycoproteins - biosynthesis; Sialoglycoproteins - isolation & purification; sialoproteins; SKELETAL DISEASES; SULFATES; Sulfates - metabolism; SULFUR 35; SULFUR COMPOUNDS; SULFUR ISOTOPES; Sulfur Radioisotopes; SYNTHESIS; TRACER TECHNIQUES; Tritium; TRITIUM COMPOUNDS; TUMOR CELLS; TYROSINE; Tyrosine - metabolism; VERTEBRATES; Purification; Radiolabelling; Rat; Gel electrophoresis; Rodentia; Glycoproteins; Glycosylation; Ion exchange chromatography; Sulfatation; Posttranslational modification; Proteins; Vertebrata; Mammalia; Cell line; Sialoproteins; Osteosarcoma; Gel filtration; Tumor
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)34119-5

The rat osteosarcoma cell line (UMR 106-01) synthesizes and secretes relatively large amounts of a sulfated glycoprotein into its culture medium (approximately 240 ng/10(6) cells/day). This glycoprotein was purified, and amino-terminal sequence analysis identified it as bone sialoprotein (BSP). [35S]Sulfate, [3H]glucosamine, and [3H]tyrosine were used as metabolic precursors to label the BSP. Sulfate esters were found on N- and O-linked oligosaccharides and on tyrosine residues, with about half of the total tyrosines in the BSP being sulfated. The proportion of 35S activity in tyrosine-O-sulfate (approximately 70%) was greater than that in N-linked (approximately 20%) and O-linked (approximately 10%) oligosaccharides. From the deduced amino acid sequence for rat BSP (Oldberg, A., Franzén, A., and Heinegård, D. (1988) J. Biol. Chem. 263, 19430-19432), the results indicate that on average approximately 12 tyrosine residues, approximately 3 N-linked, and approximately 2 O-linked oligosaccharides are sulfated/molecule. The carboxyl-terminal quarter of the BSP probably contains most, if not all, of the sulfated tyrosine residues because this region of the polypeptide contains the necessary requirements for tyrosine sulfation. Oligosaccharide analyses indicated that for every N-linked oligosaccharide on the BSP, there are also approximately 2 hexa-, approximately 5 tetra-, and approximately 2 trisaccharides O-linked to serine and threonine residues. On average, the BSP synthesized by UMR 106-01 cells would contain a total of approximately 3 N-linked and approximately 25 of the above O-linked oligosaccharides. This large number of oligosaccharides is in agreement with the known carbohydrate content (approximately 50%) of the BSP.