Separation of growth-promoting activity for human muscle cells from fetuin.
Whether the growth-promoting activity of Pedersen fetuin is due to fetuin itself or to a contaminant(s) has been a long-standing puzzle. The possibility that the growth-promoting activity of Pedersen fetuin for human muscle satellite cells (HMSC) could be caused by some other component of fetal bovine serum (FBS) that remained in the fetuin as a contaminant has been investigated. One liter of FBS was first precipitated with 50% saturated ammonium sulfate, which leaves the serum albumin in solution, and then with 25% polyethylene glycol, which leaves the fetuin in solution, to generate a fraction 50 PEG 2x that was enriched 11-fold in growth-promoting activity for HMSC, with 68% recovery of total activity. Further purification with FPLC anion exchange chromatography achieved 99-fold enrichment of the activity with 30% overall recovery. The activity is heat labile and pH sensitive, suggesting that it is of protein nature, and the size of the activity is above 70 kDa. SDS-PAGE of the most active fractions shows that they are virtually free of fetuin. Thus, although the active fractions are not homogeneous, these studies demonstrate that the growth-promoting activity for HMSC can be fully separated from fetuin.
Nie
Z
Z
Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder 80309-0347.
Jellinek
D
D
Ham
R G
RG
D000818:Animals; D001798:Blood Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D002999:Clone Cells; D003470:Culture Media; D006133:Growth Substances; D006801:Humans; D008970:Molecular Weight; D009132:Muscles / Q000166:cytology* / Q000187:drug effects; D000509:alpha-Fetoproteins / Q000302:isolation & purification / Q000494:pharmacology*
D001798:Blood Proteins; D003470:Culture Media; D006133:Growth Substances; D000509:alpha-Fetoproteins