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

Youngman Oh, Srinivasa R. Nagalla, Yoshitaka Yamanaka, Ho Seong Kim, Elizabeth Wilson, Ronald (Ron) Rosenfeld

Research output: Contribution to journalArticle

286 Citations (Scopus)

Abstract

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.

Original languageEnglish (US)
Pages (from-to)30322-30325
Number of pages4
JournalJournal of Biological Chemistry
Volume271
Issue number48
DOIs
StatePublished - 1996

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Insulin-Like Growth Factor Binding Proteins
Insulin-Like Growth Factor II
Insulin-Like Growth Factor I
Tissue
Proteins
Cells
Insulin-Like Growth Factor Binding Protein 3
Electrophoresis
Complementary DNA
Genes
Display devices
insulin-like growth factor binding protein-related protein 1
Ligands
Amino Acids
Baculoviridae
Normal Distribution
Messenger RNA
Colonic Neoplasms
Polyacrylamide Gel Electrophoresis
Amino Acid Sequence

ASJC Scopus subject areas

  • Biochemistry

Cite this

Synthesis and characterization of insulin-like growth factor-binding protein (IGFBP)-7 : Recombinant human mac25 protein specifically binds IGF-I and -II. / Oh, Youngman; Nagalla, Srinivasa R.; Yamanaka, Yoshitaka; Kim, Ho Seong; Wilson, Elizabeth; Rosenfeld, Ronald (Ron).

In: Journal of Biological Chemistry, Vol. 271, No. 48, 1996, p. 30322-30325.

Research output: Contribution to journalArticle

Oh, Youngman ; Nagalla, Srinivasa R. ; Yamanaka, Yoshitaka ; Kim, Ho Seong ; Wilson, Elizabeth ; Rosenfeld, Ronald (Ron). / Synthesis and characterization of insulin-like growth factor-binding protein (IGFBP)-7 : Recombinant human mac25 protein specifically binds IGF-I and -II. In: Journal of Biological Chemistry. 1996 ; Vol. 271, No. 48. pp. 30322-30325.
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abstract = "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.",
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AU - Wilson, Elizabeth

AU - Rosenfeld, Ronald (Ron)

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