Glycoproteins in Culture Medium: A Comparison from Cystic Fibrosis and Control Skin Fibroblasts

Thomas F. Scanlin, Judith A. Voynow, Edwin J. Thomas, Mary Catherine Glick

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16 Scopus citations

Abstract

High molecular weight glycoproteins have been partially purified by four methods from the medium after growth of human skin fibroblasts. One of these methods, precipitation by heparin, yields one major glycoprotein, Mr 220000, when examined by Polyacrylamide gel electrophoresis under denaturing conditions. The glycoprotein was labeled with l-[3H]fucose or d-[3H]glucosamine and yielded glycopeptides after digestion with Pronase. Using this method of precipitation, we made a comparison of matched cystic fibrosis (CF) and control human skin fibroblast media with and without 10% fetal calf serum, a nutritive requirement for growth. The profiles of the [3H]fucose-labeled material from media of both cell types were similar after Polyacrylamide gel electrophoresis regardless of the procedure used to obtain them. In contrast, differences were seen in the composition of the material obtained by precipitation with heparin, which suggested differences in glycosylation by CF fibroblasts. In addition, differences were noted which were related to the presence or absence of fetal calf serum in the growth medium. The analysis showed that when the CF and control materials were compared, the heparin precipitate of the CF medium without fetal calf serum contained (1) less protein and carbohydrate, (2) 30–35% less radioactivity per milligram of protein, (3) a lowered fucose content, (4) higher sialic acid content in confluency, and (5) lower turnover when expressed as counts per minute per nanomole of fucose. When the medium from the CF fibroblasts contained fetal calf serum, the heparin-precipitated material had more radioactivity and a higher ratio of fucose to other monosaccharides than that from control cells. Thus, there was either a differential response of the CF and control fibroblasts grown in the presence of 10% fetal calf serum or a differential recruitment of other glycoproteins during the precipitation of the serum-containing medium. The results are discussed in relation to previous findings of the monosaccharide content of CF fibroblast membranes and CF secretions.

Original languageEnglish (US)
Pages (from-to)491-497
Number of pages7
JournalBiochemistry
Volume21
Issue number3
DOIs
StatePublished - 1982
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biochemistry

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