Enkephalin precursors

Comb M, Seeburg PH, Adelman J, Eiden L, Herbert E (1982) Primary structure of the human Met- and Leu-enkephalin precursor and its mRNA. Nature 295 663-666. 

Metorphamide, adrenorphin, H-Tyr-Gly-Gly-Phe-Met-Arg-Arg-Val-NH2, an 8-peptide amide found in bovine brain and adrenal tissue. The sequence of metorphamide corresponds to bovine pro-enkephalin precursor-(206-213). Furthermore, metorphamide is similar in sequence to bovine adrenal medulla peptides. It shows high affinity to X opioid receptors [H. Matsuo et al.. Nature 1983, 305, 721 M. Sonders, E. Weber, J. Neurochem. 1987, 49, 671]. 

Although up to 5 mg of protein may rapidly be purified in this way, exlusion chromatography is not generally used in the neuropeptide field as many proteins of interest (e.g., enkephalin precursors) are fully amenable to purification by reverse-phase HPLC. 

In the neuropeptide field HPLC has been used successfully in recent studies on, for example, enkephalin precursors (Rossier et al., 1980 Kimura et al., 1979 Rubenstein et al, 1978 Lewis et al, 1979), B Lipotro-pin/ACTH fragments (Burbach etal, 1980), and somatostatin-like peptides (Pearson et al., 1980 Schally et al., 1980). It is most likely that these procedures may be generally applied quite successfully to neuropeptide characterization however, as yet the yield at the picomole level and applicability to all neuropeptides in crude biological samples have not been fully demonstrated. It is probable that further developmental work would be required before general application in this field is possible. 

In these pain studies, the next question to be answered concerns the origin of these increased enkephalin levels Are they biosynthesized directly or do they arise by metabolism of some larger, perhaps biologically inactive, protein Over the past few years many excellent studies on the isolation and characterization of enkephalin precursors have been reported, with reverse-phase HPLC playing a major part in the purification of both the macrolecules themselves and their enzymically generated fragments (e.g., Rossier et aL, 1980 Kimura et a/., 1979 Rubenstein et al.y 1978 Lewis et ai, 1979 Lariviere et ai, 1980 Beaumont et a/., 1980). 

Lewis, R. V., Stern, A. S., Rossier, J., Stein, S., and Undenfriend, S., 1979, Putative enkephalin precursors in bovine adrenal medulh, Biochem. Biophys. Res. Commun. 89 822-829. 

In the anterior pituitary gland (see Hormones, anteriorpituitaryhormones), both adrenocorticotropic hormones (ACTH) and the endogenous opiate hormone, P-endorphin, are synthesized from a common prohormone (2) (see Opioids,endogenous). In the adrenal medulla, five to seven copies of another opiate hormone, methionine—enkephalin (Met-enkephalin), and one copy of leucine—enkephalin (Leu-enkephalin) are synthesized from each precursor molecule (3). 

The Group III peptides come from the 256-amino acid precursor, pro-dynorphin [88402-55-5] (pro-enkephalin B). This group contains dynorphin A [80448-90-4] and B [85006-82-2] as weU as a-neoendorphin [77739-20-9] (Fig. 2), all of which can be further cleaved to form biologically active iatermediates, eg, dynorphin A g and P-neoendorphin [77739-21-0] (a-neoendorphin ) (28). The longer of these peptides are relatively basic because of the number of Lys and Arg residues. 

In addition to the weU-defined opioid systems in the central nervous system, the three opioid peptides and their precursor mRNA have also been identified in peripheral tissues. ( -Endorphin is most abundant in the pituitary, where it exists in corticotroph cells with ACTH in the anterior lobe and in melanotroph cells with MSH in the intermediate lobe (59). Enkephalin and pre-pro-enkephalin mRNA have been identified in the adrenal medulla (60) and this has been the source of material for many studies of pro-enkephalin synthesis and regulation. Pre-pro-enkephalin mRNA has also been identified in the anterior and posterior lobes of the pituitary (61). mRNA for all three opioid precursors has been identified in the reproductive system (62—64). POMC... 

Two different peptides (TRH and enkephalin, substance P and CCK) coming from different precursors. These combinations are intriguing in that TRH is excitatory yet enkephalins are inhibitory — complex postsynaptic effects can be envisaged. Substance P is excitatory and CCK acts on two receptors, A and B, with the former being the predominant CNS form. 

The classic endogenous opioid peptides are derived from one of three families of precursors proopiomelanocortin (POMC), pro-dynorphin, and pro-enkephalin. Many active opioid peptides are derived from these three, but the best known are )S-endorphin, enkephalin, and dynorphin. POMC is produced by nuclei in the hypothalamus and medulla (Khachaturian et al. 1985 Watson et al. 1978 Bloom et al. 1978). Enkephalin and dynorphin neurons are distributed to all levels of the central nervous system (Hokfelt et al. 1977 Khachaturian et al. 1983 Sar et al. 1978 Khachaturian et al. 1985). 

Endogenous opioids are peptides that are cleaved from the precursors proenkephalin, pro-opiomelanocortin, and prodynorphin. All contain the amino acid sequence of the pentapeptides [Met]- or [Leuj-enkephalin (A). The effects of the opioids can be abolished by antagonists (e.g., naloxone A), with the exception of buprenorphine. 

Precursor peptides (1)pro-endomorphin (2)pro-opiomelanocortin, <3)pro-enkephalin, (4)pro-dynorphin, (5)pro-nociceptin ( presumed to exist). 

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