Purification and characterization of a γ-melanotropin precursor from frozen human pituitary glands

Abstract

A new melanocyte-stimulating peptide was isolated from acid extracts of frozen human pituitary glands by salt/ethanol fractionation, Sephadex G-75 gel filtration and DEAE- and carboxymethyl-cellulose ion-exchange chromatography. The peptide is glycosylated, has an N-terminal tryptophan residue and an apparent MW of 16,000 as estimated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Its amino acid analysis closely resembles residues Trp-105 to Gln-29 predicted for the common precursor protein of bovine ACTH and .beta.-lipotropin. This fragment is expected to have melanotropin activity due to the tetrapeptide -His-Phe-Arg-Trp- (residues -51 to -48) of the predicted sequence of the common precursor. It has a molar potency of 1 .times. 10-5 relative to .alpha.-melanotropin in the frog skin bioassay. These characteristics are consistent with the isolated melanotropin peptide being a non-corticortropin, non-lipotropin peptide of the human common precursor protein of ACTH and lipotropin. The peptide neither potentiates the adrenal weight-maintenance activity of ACTH-(1-24)-tetracosapeptide when administered to hypophysectomized rats, nor stimulates release of non-esterified fatty acids from isolated rat epididymal cells. A 2nd N-terminal-tryptophan glycopeptide was also isolated, which had an amino-acid composition similar to that predicted for the bovine common precursor protein, residues Trp-105 to Gly-35.