Structure of the Mr 140,000 Growth Hormone-Dependent Insulin-Like Growth Factor Binding Protein Complex: Determination by Reconstitution and Affinity-Labeling

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 86; no. 18; pp. 6898 - 6902
• Main Authors: Baxter, Robert C., Martin, Janet L.
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01.09.1989; National Acad Sciences
• Subjects: 550201 - Biochemistry- Tracer Techniques; Acromegaly - blood; Adult; AFFINITY; Affinity Labels - metabolism; Amniotic fluid; Analytical, structural and metabolic biochemistry; ANIMALS; Antiserum; AUTORADIOGRAPHY; BASIC BIOLOGICAL SCIENCES; BETA DECAY RADIOISOTOPES; BIOASSAY; Biological and medical sciences; Carrier Proteins - drug effects; Carrier Proteins - isolation & purification; Carrier Proteins - metabolism; COMPLEXES; DAYS LIVING RADIOISOTOPES; ELECTRON CAPTURE RADIOISOTOPES; ELECTROPHORESIS; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Gels; GLYCOPROTEINS; GROWTH FACTORS; Growth Hormone - pharmacology; Growth hormones; HORMONES; Humans; IMMUNOASSAY; INSULIN; Insulin like growth factor binding proteins; Insulin-Like Growth Factor I - metabolism; Insulin-Like Growth Factor II - metabolism; INTERMEDIATE MASS NUCLEI; IODINE 125; IODINE ISOTOPES; Iodine Radioisotopes; ISOTOPES; Macromolecular Substances; MAMMALS; MAN; MITOGENS; Models, Structural; MOLECULAR STRUCTURE; MOLECULAR WEIGHT; NUCLEI; ODD-EVEN NUCLEI; ORGANIC COMPOUNDS; PEPTIDE HORMONES; Phosphates; PRIMATES; Product labeling; Protein hormones. Growth factors. Cytokines; PROTEINS; RADIOISOTOPES; Somatomedins - metabolism; VERTEBRATES; Human; Radiolabelling; Somatomedin C; Ternary complex; Gel electrophoresis; Insulin like growth factor 2; Chromatography; Autoradiography; Binding protein; Binding capacity; Reconstituted system; Serum; Models; Growth factor
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.86.18.6898

To determine the structure of the high molecular weight, growth hormone-dependent complex between the insulin-like growth factors (IGF-I and IGF-II) and their binding proteins in human serum, we have reconstituted the complex from its purified component proteins and analyzed it by gel electrophoresis and autoradiography after covalent cross-linking. The proteins tested in reconstitution mixtures were an acid-labile Mr 84,000-86,000 glycoprotein doublet (α subunit), an acid-stable Mr 47,000-53,000 glycoprotein doublet with IGF-binding activity (BP-53 or β subunit), and IGF-I or IGF-II (γ subunit). In incubations containing any one of the three subunits 125I-labeled and the other two unlabeled, identical 125I-labeled α -β -γ complexes of Mr 140,000 were formed. Minor bands of Mr 120,000 and 90,000 were also seen, thought to represent a partially deglycosylated form of the α -β -γ complex, and an α -γ complex arising as a cross-linking artifact. When serum samples from subjects of various growth hormone status were affinity-labeled with IGF-II tracer, a growth hormone-dependent Mr 140,000 band was seen, corresponding to the reconstituted α -β -γ complex. Other growth hormone-dependent labeled bands, of Mr 90,000 (corresponding to α -γ ), Mr 55,000-60,000 (corresponding to labeled β -subunit doublet), and smaller bands of Mr 38,000, 28,000, and 23,000-25,000 (corresponding to labeled β -subunit degradation products), were also seen in the affinity-labeled serum samples and in the complex reconstituted from pure proteins. All were immunoprecipitable with an anti-BP-53 antiserum. We conclude that the growth hormone-dependent Mr 140,000 IGF-binding protein complex in human serum has three components: the α (acid-labile) subunit, the β (binding) subunit, and the γ (growth factor) subunit.