Further characterization of the major forms of reptile beta-endorphin

Abstract

Biosynthetically labeled reptile intermediate pituitary beta-endorphin-sized material was fractionated by SP-Sephadex ion exchange chromatography into two major opiate-active forms which eluted at 0.28 M NaCl and 0.32 M NaCl, respectively; the 0.32 M form of reptile β-endorphin ( mw=3500 ), serves as the precursor for the 0.28 M form of reptile β-endorphin ( mw=3200 ), (Dores and Surprenant, 1983). Analysis of tryptic digests of these reptile β-endorphins by paper electrophoresis at pH 3.5 and gel filtration on a Sephadex G-15 column indicated that there are two tyrosine residues, two arginine residues and one methionine residue in reptile β-endorphin. Furthermore, the NH 2-terminal tryptic peptide of both reptile β-endorphins is approximately nine amino acids in size and contains tyrosine, methionine and arginine. Analyses of chymotryptic/protease digests of the [ 3H]tyrosine-labeled NH 2-terminal tryptic peptide analyzed by descending paper chromatography revealed that the NH 2-terminal tyrosine of reptile β-endorphin is not α-N-acetylated. A second tyrosine-containing tryptic peptide was detected in the COOH-terminal region of reptile β-endorphin; however this tryptic peptide differs in the two forms of reptile β-endorphin in terms of size and net charge at pH 3.5. These differences account for the apparent molecular weight differences and distinct ion exchange properties of the 0.28 M and 0.32 M forms of reptile β-endorphin. Thus in the reptile intermediate pituitary the principal post-translational mechanism for modifying β-endorphin is COOH-terminal proteolytic cleavage.