In:
Biochemistry and Cell Biology, Canadian Science Publishing, Vol. 69, No. 8 ( 1991-08-01), p. 523-530
Abstract:
Following incubation of UMR-106 cells for 48 h in the presence of [ 3 H]glucosamine and [ 35 S]sulfate, the newly synthesized anionic glycoconjugates were isolated from the culture medium by cetylpyridinium chloride/ethanol precipitation and further separated by DEAE-Sephacel chromatography into two radiolabelled fractions, a major component, UM I, and a minor component, UM II. UM I appeared to be homogeneous as shown by Sepharose CL-4B chromatography under dissociative conditions, and SDS-polyacrylamide gel electrophoresis. It showed a molecular mass of approximately 93 kDa on 4–15% gels. UM I was partially degraded by brief treatment with trypsin, releasing a small, terminal peptide that contained 47.6% of 35 S but no 3 H. Treatment of UM I with neuraminidase and 0.1 N H 2 SO 4 (1 h at 80 °C), respectively, released 27% 3 H and 38.4% 3 H plus 41% 35 S, suggesting the presence of a significant number of sialic acid residues, as shown by Sephadex G-50 chromatography of the digests. Amino acid analysis showed that the UM I glycoconjugate was rich in acidic amino acids (12.6% aspartic acid and 21.2% glutamic acid residues) and its N-terminal sequence was Phe-Ser-Met-Lys-Asn-Phe-, which is identical to the published N-terminal amino acid sequence of rat bone sialoprotein II. Keratanase treatment of UM I released 26% of the incorporated radioactivity, suggesting the presence of keratan sulfate chains. UM II contained a chondroitinase ABC-sensitive proteoglycan.Key words: UMR-106 cells, anionic glycoconjugates, bone sialoprotein II.
Type of Medium:
Online Resource
ISSN:
0829-8211
,
1208-6002
Language:
English
Publisher:
Canadian Science Publishing
Publication Date:
1991
SSG:
12
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