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The major N-terminal acetylated endorphin of the pars intermedia of Xenopus laevis was purified and submitted to fast-atom bombardment tandem mass spectroscopy. The collisionally induced dissociation MS/MS spectrum of the [M+H]+ ion revealed sufficient fragment ions to determine unambiguously the identity of the peptide as alpha,N-acetyl beta-endorphin [1-8], the sequence of which was predicted on the basis of the nucleotide sequence of Xenopus POMC cDNA. The determination was confirmed by showing that the synthetic peptide of this structure had identical FAB tandem mass spectrometric characteristics as the endogenous endorphin. We conclude that alpha,N-acetyl beta-endorphin [1-8] is the terminal product of processing of endorphins in the melanotrope cell of Xenopus laevis.

High affinity binding sites for beta-endorphin1-31 (beta-EP) have been observed on transformed mononuclear cells such as the human U937 monocyte-like cell line and the murine EL4-thymoma line, and on normal murine splenocytes. Binding of beta-EP at these sites is resistant to competition by naloxone and other opiate receptor ligands but sensitive to N-acetyl-beta-endorphin1-31 (N-Ac), cations and GTP-gamma-sulfate. Thus, the following studies were done to determine the functional significance of binding beta-EP and N-Ac. beta-EP suppressed phytohemagglutinin (PHA)-stimulated [3H]thymidine uptake in a dose-dependent, naloxone-insensitive fashion. beta-Endorphin1-27, (des)-tyrosine beta-endorphin2-31, or N-Ac failed to duplicate the suppressive effect of beta-EP. However, N-Ac, which is equipotent to beta-EP at displacing 125I-beta-EP bound to murine splenocytes or U937 cells, antagonized the suppressive effect of beta-EP. Taken together with previous binding studies, the present observations suggest that beta-EP effects receptor-mediated responses on normal immunocytes that do not depend on the activation of neuronal-like opiate receptors which are naloxone-sensitive. N-Ac, which shows minimal binding to such brain opiate receptors, is a potent functional antagonist of the naloxone-insensitive immunocyte receptor for beta-EP.