Isolation and characterization of corticotropin- and melanotropin-related peptides from the neurointermediary lobe of the rat pituitary by reversed-phase liquid chromatography

Abstract

A novel procedure utilizing reversed-phase high-performance liquid chromatography for the extraction and purification of peptides from biological tissues was applied to the isolation of corticotropin-like intermediary lobe peptide (CLIP) and .alpha.-melanocyte-stimulating hormone (.alpha.-MSH) from the neurointermediary lobe of the rat pituitary. The isolation and characterization of 2 major forms of CLIP and 2 major forms of .alpha.-MSH are described. The isolated peptides were identified using enzymatic digestions and peptide mapping. The main form of CLIP is a peptide which was modified by phosphorylation of the Ser residue at position 31. This is the 1st peptide of endocrine origin reported to be modified in such a manner. A nonphosphorylated form of CLIP was also present at lower concentrations. The main form of .alpha.-MSH was N,-O-diacetyl-.alpha.-MSH, with the more familiar mono-N-acetyl-.alpha.-MSH present to a much smaller extent. Thus, in the rat neurointermediary lobe, the 2 main ACTH-related peptides present are mostly in modified forms, which are the result of posttranslational modifications. It is only by the use of methodology such as that previously described that small alterations in peptide structure may be identified.