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Localization of Endo‐Oligopeptidase (EC 3.4.22.19) in the Rat Nervous Tissue
Author(s) -
Oliveira Elisabeth S.,
Leite Paulo E. P.,
Spillantini Maria G.,
Camargo Antonio C. M.,
Hunt Stephen P.
Publication year - 1990
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.1471-4159.1990.tb03113.x
Subject(s) - oligopeptidase , neocortex , striatum , cerebellum , synaptosome , biochemistry , chemistry , central nervous system , enzyme , microbiology and biotechnology , biology , membrane , endocrinology , neuroscience , dopamine
The subcellular and regional distribution of endooligopeptidase (EC 3.4.22.19), an enzyme capable of generating enkephalin by single cleavage from enkephalin‐containing peptides, was determined by an enzymatic assay using metorphamide and by immunochemical techniques in the CNS of the rat. The rat CNS contains a membrane‐associated form of endo‐oligopeptidase, an enzyme predominantly associated with the soluble fraction of brain homogenates. Sub‐cellular fractionation showed that ∼17% of the total activity of the enzyme is associated with membrane fractions including synaptosomes. Synaptosomal membranes were prepared from neocortex, striatum, hypothalamus, medulla, spinal cord, and cerebellum. The amount of EC 3.4.22.19 activity solubilized by 3‐([3‐cholamidopropyl]dimethylammonio)‐1‐propanesulfonate from synaptosomal membranes was similar in neocortex, striatum, and hypothalamus, being three‐ to 10‐fold greater than in spinal cord, cerebellum, and medulla. A polyclonal antibody exhibiting high affinity for endo‐oligopeptidase was raised in rabbits against the purified rat brain enzyme and used to localize endo‐oligopeptidase by Western blotting and by immunoperoxidase techniques. A strong band corresponding to the M r of EC 3.4.22.19 was found in solubilized proteins obtained from synaptosomal membranes prepared from hypothalamus, neocortex, and striatum when subjected to Western blotting. The immunohistochemical localization of endo‐oligopeptidase indicated that the immunoreactivity was confined to gray matter in regions known to be rich in peptide‐containing neurons such as the striatum. In the cerebellum, a region poor in peptides, no staining could be detected. The nonuniform distribution of endo‐oligopeptidase in rat brain suggests a role in neurotransmitter processing in the CNS.