A general goal of this proposed project is to investigate the function of beta-endorphin (Beta-EP) and related psychopharmacologically significant peptides at the molecular level.
The specific aims are as follows: (a) To investigate on the isolation and characterization of the opioid and non-opioid beta-EP binding proteins (receptors) from membrane preparations of rat brains, mouse neuroblastoma x glioma hybrid cells (NG108-15 and N18TG2), human glioblastoma SF126 cells and human neuroblastoma IMR-32 cells using a photoaffinity reagent 2-nitro-4-azidophenylsulfenyl chloride specific to label tryptophan residue; (b) To synthesize two analogs ([Trp-4]-beta-h-EP; [D-Trp-4]-beta-h-EP) by solid-phase method to use for photolabel probes of the opioid domains in the beta-h-EP molecule. Synthetic [Trp-27]-beta-h-EP will be used for photolabel probes of non-opioid binding sites; (c) To further characterize biologically the naturally occurring beta-EP-inhibiting peptide [beta-EIP, beta-EP-(1-27)] and the synthetic analogs with inhibiting beta-EP analgesia property by radioreceptor binding assay using rat brain membrane preparations and tritiated dihydromorphine, [D-Ala-2, D-Leu-5]-enkephalin, ethylketocyclazocine, beta-EP and diprenorphine as primary ligands as well as by bioassays using guinea pig ileum, rat, mouse and rabbit vas deferens preparations; (d) to test inhibiting activities of beta-h-EP-(1-27) in other biological systems such as prolactin-releasing activity of beta-EP; and (e) to isolate, synthesize and characterize beta-EP from elephant pituitary glands.
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