Fetuin and alpha-2HS glycoprotein induce alkaline phosphatase in epiphyseal growth plate chondrocytes.
A previously described chondrocyte alkaline phosphatase induction factor (CAP-IF) for chicken epiphyseal growth plate chondrocytes has been purified to SDS-PAGE homogeneity from fetal bovine serum by ammonium sulfate precipitation and by dye-ligand affinity (Affi-Gel Blue and Reactive Green-19 agarose) and hydroxyapatite column chromatographies. As determined by immunoprecipitation of [35S]methionine-labeled cellular proteins after 3 day treatment, this highly purified CAP-IF increases the level of AP and certain other membrane proteins 2- to 3-fold over control values. The pure protein of apparent 64.5 kDa molecular weight has been identified as fetuin by N-terminal amino acid sequencing. This was confirmed by the finding that high alkaline phosphatase (AP)-inducing activity is present in fetuin prepared by the Spiro method. However, fetuins prepared by the Pedersen or Deutsch procedures are inactive. At least half of the CAP-IF activity of fetuin was irreversibly destroyed by treatment with EDTA and addition of Zn2+ did not reactivate the EDTA-treated fetuin. Ascorbate synergistically enhanced the effect of fetuin on chondrocyte AP activity by over 8-fold during 3 day exposure. Because of the very high homology between fetuin and the A-chain of alpha 2-HS glycoprotein, we also tested and found that alpha 2HS glycoproteins from human serum and bovine bone are both strong AP inducers. Our findings suggest that the AP-inducing activity resides in a labile, cystatin/Zn(2+)-binding domain common to these related serum glycoproteins. These proteins appear to play a role in enhancing AP expression in normal growth plate cartilage differentiation.
Ishikawa
Y
Y
Department of Chemistry, University of South Carolina, Columbia 29208.
Wu
L N
LN
Valhmu
W B
WB
Wuthier
R E
RE
D000469:Alkaline Phosphatase / Q000096:biosynthesis*; D000595:Amino Acid Sequence; D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology; D002645:Chickens; D004357:Drug Synergism; D004790:Enzyme Induction; D005353:Fibronectins / Q000494:pharmacology*; D006132:Growth Plate / Q000201:enzymology*; D008565:Membrane Proteins / Q000096:biosynthesis; D008969:Molecular Sequence Data; D010716:Phosphatidylinositols / Q000378:metabolism; D000509:alpha-Fetoproteins / Q000302:isolation & purification / Q000494:pharmacology*
D005353:Fibronectins; D008565:Membrane Proteins; D010716:Phosphatidylinositols; D000509:alpha-Fetoproteins; D000469:Alkaline Phosphatase; D001205:Ascorbic Acid