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>Characterization of the increased biological potency in BALB/C 3T3 cells of two analogs of human insulinlike growth factor I which have reduced affinity for the 28 K cell‐derived binding protein
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Characterization of the increased biological potency in BALB/C 3T3 cells of two analogs of human insulinlike growth factor I which have reduced affinity for the 28 K cell‐derived binding protein
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机译：Characterization of the increased biological potency in BALB/C 3T3 cells of two analogs of human insulinlike growth factor I which have reduced affinity for the 28 K cell‐derived binding protein
AbstractWe have characterized the biological activity of two analogs of insulinlike growth factor I (IGF I) which have significantly reduced affinity for the soluble 28 K binding proteins which are secreted by various cell types. The analogs, which were made by site‐directed mutagenesis of a synthetic gene encoding for IGF I, are [Gln 3, Ala 4, Tyr 15, Leu 16] IGF I and an analog in which the first 16 amino acids of IGF I were replaced with the first 17 amino acids of insulin (B‐chain mutant). These two peptides have 100‐fold and>1,000‐fold lower affinity, respectively, than IGF I for the 28 K binding protein present in the conditioned medium of two cell types, the clonal rat vascular smooth muscle line A10, and BALB/C 3T3 cells. The 28 K protein secreted by BALB/C 3T3 cells has fivefold‐lower apparent affinity for both IGF I and [Gln 3, Ala 4, Tyr 15, Leu 16] IGF I than does the 28 K protein secreted by A10 cells. Conditioned medium from these two cells types has similar amounts of unoccupied 28 K protein as evidenced by the ability of125I‐IGF I to specifically bind to and be covalently bound to the protein after treatment with the bifunctional cross‐linking reagent disuccinimidyl suberate.In the presence of 0.1% calf serum, IGF I and [Gln 3, Ala 4, Tyr 15, Leu 16] IGF I stimulate DNA synthesis in A10 cells with ED50= 0.4 nM, and in BALB/C 3T3 cells with ED50= 10 nM and 1.3 nM, respectively. Thus, these peptides are equipotent in A10 cells, but the mutant peptide is ten times more active than IGF I in BALB/C 3T3 cells. A10 cells can be made ten times less sensitive to IGF I by performing the incubation in the presence of conditioned media from BALB/C 3T3 cells but not from A10 cell?. The activity of [Gln 3, Ala 4, Tyr 15, Leu 16] IGF I is not altered under these conditions. Thus, the conditioned media, which contain 28 K proteins secreted by A10 cells and BALB/C 3T3 cells, have different effects on the biological action of IGF I. These data suggest that the 28 K binding proteins can have important effects on the sensitivity of tissues to IGF I and that the B‐chain mutant and [Gln 3, Ala 4, Tyr 15, Leu 16]IGF I will be useful in assessing the biological role of
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