An insulin-like growth factor-binding protein purified from medium conditioned by a human lung fibroblast cell line (He[39]L) has a novel N-terminal sequence.
We have identified a novel insulin-like growth factor (IGF)-binding protein secreted into the culture medium of a human lung fibroblast cell line (He[39]L). This binding protein was purified by S-Sepharose cation exchange chromatography, IGF affinity chromatography and reverse-phase high-performance liquid chromatography. Analysis of the He[39]L-binding protein on silver-stained polyacrylamide gels and by ligand blotting showed that it had a molecular weight of 32 kDa under non-reducing conditions. In charcoal competition binding assays, IGF-II competed at lower concentrations than IGF-I for binding to the He[39]L-binding protein and the more potent form of IGF-I (des-(1-3)-IGF-I) was not bound. This is a similar IGF binding pattern to that of the bovine IGF-binding protein-2 (bIGFBP-2). However, immunoblotting with an antibody to bIGFBP-2 demonstrated that the He[39]L-binding protein is not immunochemically related to bIGFBP-2. It is a glycosylated protein, having N-acetyl-glucosaminyl sugars detected by wheatgerm agglutinin affinity chromatography. Of the first 15 N-terminal amino acids of the He[39]L-binding protein, 13 are identical to the 15 amino acid sequence of a recently sequenced cerebrospinal fluid-binding protein. However, the total He[39]L-binding protein sequence (25 amino acids) showed no homology to other previously sequenced binding proteins (IGFBP-1, IGFBP-2 and IGFBP-3). We conclude that the He[39]L-binding protein is a novel binding protein.
Department of Biochemistry, University of Adelaide, South Australia.
B
B
Forbes
F J
FJ
Ballard
J C
JC
Wallace
D000595:Amino Acid Sequence; D015152:Blotting, Northern; D002352:Carrier Proteins / Q000302:isolation & purification*; D002460:Cell Line; D002846:Chromatography, Affinity; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D018969:Insulin-Like Growth Factor Binding Proteins; D008168:Lung / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight
D002352:Carrier Proteins; D018969:Insulin-Like Growth Factor Binding Proteins