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Isolation and Characterization of Opioid Peptides from Rabbit Cerebellum
Author(s) -
Madden John,
Evans Christopher J.,
Tyler Andrew N.,
Esch Frederick S.,
Böhlen Peter,
Makk George,
Weber Eckard
Publication year - 1991
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1991.tb03448.x
Subject(s) - proenkephalin , opioid peptide , dynorphin , opioid , enkephalin , chemistry , peptide , cerebellum , peptide sequence , radioimmunoassay , receptor , dynorphin a , leu enkephalin , opioid receptor , biochemistry , endocrinology , biology , gene
Abstract: The rabbit cerebellum has been shown to contain significant quantities of opioid receptors consisting of both γ‐ and k ‐subtypes. To determine the nature of the endogenous opioid ligands in this tissue, extracts from rabbit cerebellum were separated by various chromatography techniques and fractions were assayed initially for opioid peptides with a radioimmunoassay capable of detecting all peptides with an amino‐terminal Tyr‐Gly‐Gly‐Phe sequence. This sequence is common to all mammalian opioid peptides and is critical for recognition by all known opioid receptors. Each of the three immunoreactive opioid peptide peaks detected was purified to homogeneity and subjected to amino acid composition and sequence analysis. One peak was analyzed further by mass spectrometry. This identified the major opioid peptides in the cerebellum as [Met 5 ]enkephalin, [Leu 5 ]enkephalin, and heptapeptide [Met 5 ]enkephalyl‐Arg 6 ‐Phe 7 . The comprehensiveness of this initial detection scheme in identifying biologically active opioid peptides was substantiated through subsequent analysis. Using specific radioimmunoassays for representative opioid peptides of the three opioid systems currently known, no other peptides of either the proenkephalin, proopiomelanocortin, or prodynorphin series were detected in any appreciable amounts. Collectively, these results are consistent with the position that rabbit cerebellar opioids are derived from proenkephalin. However, given that no appreciable quantities of either [Met 5 ]enkephalyl‐Arg 6 ‐Arg 7 ‐Val 8 ‐NH 2 (metorphamide) or [Met 5 ]enkephalyl‐Arg 6 ‐Gly 7 ‐Leu 8 were detected suggests that rabbit proenkephalin may have a slightly altered sequence and/or is differentially processed relative to other mammalian species studied.