Detection of a novel immunoreactive endomorphin 2-like peptide in rat brain extracts.
To pursue further the possible de novo biosynthetic pathway of endomorphins in rat brain we raised antibodies to endomorphin-2 conjugate in rabbits. Antiserum R1 recognized endomorphin-2 with good selectivity as compared to endomorphin-1 with a median detection value of 65.5+/-7.5 pg/tube (n=7), whereas R4 antiserum recognized both endomorphins with similar sensitivity. Neither antisera recognized YP-related di- or tripeptides or YGGF-related opioid sequences (enkephalins, beta-endorphin, dynorphin). Using the same rat brain extraction-RP-HPLC-gradient separation paradigm as previously, antisera detected 144.6+/-40.0 (n=3) pg/g wet brain weight endomorphin-2-like immunoreactivity in the fraction corresponding to standard endomorphin-2 retention time and also in the fraction matching endomorphin-2-OH standard retention time (179.1+/-30.1 pg/g). Since R1 failed to recognize authentic endomorphin-2-OH, the second immunoreactive species must be different from both endomorphin-2 and endomorphin-2-OH. Possible biosynthetic intermediates to endomorphins, synthetic YPFFG and YPWFG had retention times close to the parent endomorphin standards in RP-HPLC gradient separation profile. The former was a mu-opioid receptor agonist of medium potency in the in vitro assays (rat brain RBA&GTP gamma S binding and mouse vas deferens), whereas the latter was a weak mu-opioid receptor agonist with a significant delta-opioid receptorial action as well and a definite indication of partial agonism.
Szemenyei
Erzsébet
E
HAS Biological Research Center, Institute of Biochemistry, P.O.B. 521, H-6701, Szeged, Hungary.
Barna
István
I
Mergl
Zsuzsa
Z
Keresztes
Attila
A
Darula
Zsuzsanna
Z
Kató
Erzsébet
E
Tóth
Géza
G
Rónai
András Z
AZ
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000276:immunology*; D002851:Chromatography, High Pressure Liquid; D004399:Dynorphins / Q000276:immunology; D004745:Enkephalins / Q000276:immunology; D007106:Immune Sera / Q000276:immunology; D008297:Male; D051379:Mice; D009292:Narcotic Antagonists / Q000276:immunology; D009842:Oligopeptides / Q000276:immunology*; D010455:Peptides / Q000276:immunology* / Q000302:isolation & purification; D011817:Rabbits; D011863:Radioimmunoassay / Q000379:methods; D051381:Rats; D017208:Rats, Wistar; D001615:beta-Endorphin / Q000276:immunology
D004745:Enkephalins; D007106:Immune Sera; D009292:Narcotic Antagonists; D009842:Oligopeptides; D010455:Peptides; C105294:endomorphin 2; D001615:beta-Endorphin; D004399:Dynorphins