Synthesis and characterization of insulin-like growth factor-binding protein (IGFBP)-7. Recombinant human mac25 protein specifically binds IGF-I and -II

• Published in: The Journal of biological chemistry Vol. 271; no. 48; pp. 30322 - 30325
• Main Authors: Oh, Y, Nagalla, S R, Yamanaka, Y, Kim, H S, Wilson, E, Rosenfeld, R G
• Format: Journal Article
• Language: English
• Published: United States 29-11-1996
• Subjects: Gene Expression; Humans; Insulin-Like Growth Factor Binding Proteins - biosynthesis; Insulin-Like Growth Factor Binding Proteins - chemistry; Insulin-Like Growth Factor I - metabolism; Insulin-Like Growth Factor II - metabolism; Peptides - metabolism; Recombinant Proteins; RNA, Messenger - genetics; Tissue Distribution
• ISSN: 0021-9258
• DOI: 10.1074/jbc.271.48.30322

The mac25 cDNA was originally cloned from leptomeningial cells and subsequently reisolated through differential display as a sequence preferentially expressed in senescent human mammary epithelial cells. The deduced amino acid sequence of the human mac25 propeptide shares a 20-25% identity to human insulin-like growth factor-binding proteins (IGFBPs), suggesting that mac25 could be another member of the IGFBP family. In the present study, we have generated recombinant human mac25 (rh-mac25) in a baculovirus expression system and assessed its affinity for IGFs and have evaluated the pattern of expression of the mac25 gene in human tissues. Binding of 125I-IGF-I and 125I-IGF-II to rh-mac25 was demonstrated by Western ligand blotting after nondenaturing polyacrylamide gel electrophoresis and by affinity cross-linking with as little as 2 nM rh-mac25. Specificity of rh-mac25 binding to 125I-IGFs was demonstrated by competition for rh-mac25 binding with unlabeled IGFs, but not with [QAYLL]IGF-II analog, which has 100-fold less affinity for IGFBPs. In comparison with IGFBP-3, rh-mac25 has at least a 5-6-fold lower affinity for IGF-I and 20-25-fold lower affinity for IGF-II. mac25 mRNA was detectable in a wide range of normal human tissues, with decreased expression in breast, prostate, colon, and lung cancer cell lines. In conclusion, mac25 specifically binds IGFs and constitutes a new member of the IGFBP family, IGFBP-7. Its wider distribution in normal tissue and lower expression in several cancer cells indicate that IGFBP-7 may function as a growth-suppressing factor, as well as an IGF-binding protein.