Purification and properties of a novel insulin-like growth factor-II binding protein from transformed human fibroblasts.
We have purified an insulin-like growth factor (IGF) binding protein from culture medium conditioned by the SV40-transformed human fibroblast line AG 2804. This protein (TFBP), of apparent Mr = 34,000 on gel electrophoresis, binds IGF-II with an affinity constant of 3 x 10(11) liters/mol, 100-fold higher than its affinity for IGF-I. Amino-terminal sequencing indicated no structural relationship to any previously characterized IGF binding protein or receptor. Like a protein in cerebrospinal fluid with selective affinity for IGF-II, TFBP does not bind IGF-I lacking the amino-terminal tripeptide. In contrast with binding proteins produced by normal human fibroblasts, TFBP does not cross-react in radioimmunoassays for the plasma protein IGFBP-3 or the amniotic fluid protein IGFBP-1; however, after affinity labeling with IGF-II, it is immunoprecipitable by an anti-IGFBP-3 antiserum. The cerebrospinal fluid binding protein is not precipitable by this antiserum and appears smaller than TFBP. TFBP reacts with wheat germ agglutinin, but not with concanavalin A, or with the acid-labile subunit of the high molecular weight serum IGF binding protein complex. Furthermore, since RNA from transformed fibroblasts does not hybridize with an IGFBP-3 cDNA probe, TFBP is not derived from the IGFBP-3 gene. We conclude that, although TFBP appears to share an epitope with IGFBP-3, it is distinctly different from any previously described IGF binding protein.
Martin
J L
JL
Department of Endocrinology, Royal Prince Alfred Hospital, Camperdown, NSW, Australia.
Willetts
K E
KE
Baxter
R C
RC
D000595:Amino Acid Sequence; D001667:Binding, Competitive; D015152:Blotting, Northern; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007335:Insulin-Like Growth Factor II / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011956:Receptors, Cell Surface / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D017451:Receptors, Somatomedin; D013539:Simian virus 40 / Q000235:genetics*; D013002:Somatomedins / Q000378:metabolism*
D011956:Receptors, Cell Surface; D017451:Receptors, Somatomedin; D013002:Somatomedins; D007335:Insulin-Like Growth Factor II