Partial purification of a specific inhibitor of the insulin-like growth factors by reversed-phase high-performance liquid chromatography.
We report here preliminary data using reversed-phase high-performance liquid chromatography for the purification of a specific inhibitor (a molecular weight 16,000-18,000 protein) of the insulin-like growth factor (IGF) or somatomedin family. Crude inhibitor prepared from Cohn fraction IV-1 of human serum was first partially purified using an IGF/CH-Sepharose 4B affinity column. Following elution of the bound inhibitor and resuspension in 0.1% aqueous trifluoroacetic acid (mobile phase A), it was injected (100 microliter; 2.0 mg protein) onto a Brownlee Aquapore RP-300 column. Application of a linear gradient from 0% to 100% mobile phase B (45% isopropanol-0.1% trifluoroacetic acid) resulted in elution of two peaks of inhibitor activity between 31% and 34% isopropanol associated with a major homogeneous protein peak and a minor heterogeneous protein peak. No inhibitor was recovered when an acetonitrile gradient was used instead of isopropanol, indicating that the inhibitor is very hydrophobic. These data suggest that high-performance liquid chromatography offers a simple procedure for the potential purification of IGF inhibitor(s) from normal human serum.
D001798:Blood Proteins / Q000302:isolation & purification*; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002846:Chromatography, Affinity / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006801:Humans; D007328:Insulin / Q000302:isolation & purification; D007329:Insulin Antagonists; D010455:Peptides / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D013002:Somatomedins / Q000037:antagonists & inhibitors* / Q000302:isolation & purification
D001798:Blood Proteins; D007328:Insulin; D007329:Insulin Antagonists; D010455:Peptides; D013002:Somatomedins