In this paper we report the presence of a particulate pyroglutamate aminopeptidase in guinea-pig brain tissue. This activity appears to reside in the synaptosomal membrane and could be released from the membrane by treatment with papain or Triton X-100. By contrast with a previously described broad-specificity soluble pyroglutamate aminopeptidase from guinea-pig brain tissue, the enzyme released from the synaptosomal membrane preparation removed pyroglutamic acid from thyroliberin, acid thyroliberin and less than Glu-His-Gly alone of the peptides tested. Unlike the soluble tissue enzyme the present enzyme was inhibited by the presence of EDTA and the activity released from synaptosomal membranes by papain was found to have a relative molecular mass of 230 000, almost one order of magnitude greater than that reported for the soluble enzyme.
"pyroglutamyl peptidase II (PPII), that possesses a stringent specificity for the peptide  and when inhibited, the recovery of in vitro released TRH is increased . PPII is an ectoenzyme  enriched in brain and present also in adenohypophysis  where it is regulated in different conditions that involve TRH such as hyperthyroidism   or estrous cycle ; in vitro, TRH or the activation of protein kinase C can downregulate adenohypophyseal PPII activity . "
[Show abstract][Hide abstract] ABSTRACT: Pyroglutamyl peptidase II (PPII) is a neuronal ectoenzyme responsible for thyrotropin releasing hormone (TRH) degradation at the synaptic cleft. PPII, heterogeneously distributed in different brain regions and adenohypophysis, is regulated under various endocrine conditions where TRH is involved in thyrotropin or prolactin regulation but only at the adenohypophyseal level. TRH can downregulate PPII activity in cultured adenohypophyseal cells. TRH present in extrahypothalamic brain areas has been postulated to serve as a neuromodulator and levels of this peptide increase in amygdala, hippocampus and cortex after electrical stimulation (kindling or electroshock). To study whether brain PPII could be regulated in conditions that stimulate TRHergic neurons, TRH and PPII activity were determined during the development of amygdaloid kindling in the rat. TRH levels increased from stage II to V in amygdala and hippocampus in the ipsi-and contralateral side to stimulation. In n. accumbens a decrease, compared to sham was observed at stage II, but levels raised through stage V. In contrast, PPII activity was increased at stage II, in amygdala of both sides and in hippocampus, frontal cortex, n. accumbens and hypothalamus of the contralateral side; levels decreased at stage V to sham values in most structures (except amygdala and hippocampus where the activity was 30% below controls). These results suggest that PPII activity in the central nervous system can be regulated in conditions known to affect TRHergic neurons.
Brain Research 08/2013; 679(1):144-150. · 2.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A pyroglutamate aminopeptidase activity, distinct from that of cytoplasm, was released from a synaptosomal membrane preparation of guinea-pig brain by papain treatment. This activity was further purified 3560-fold relative to the homogenate with a yield of 17% by a procedure involving gel filtration chromatography, calcium phosphate cellulose chromatography and hydrophobic interaction chromatography on phenyl-Sepharose CL-4B. The purified synaptosomal pyroglutamate aminopeptidase hydrolysed only thyroliberin, acid-thyroliberin, the luliberin N-terminal tripeptide (Glp-His-Trp) and, only slightly, Glp-His-Gly. No hydrolysis was observed with dipeptides containing N-terminal pyroglutamic acid (Glp) or with pyroglutamyl peptides containing more than three amino acids. A Km value of 40 microM was recorded when thyroliberin was used as substrate; however, luliberin was found to inhibit the hydrolysis of thyroliberin competitively with a Ki value of 20 microM.
European Journal of Biochemistry 08/1985; 150(1):47-52. DOI:10.1111/j.1432-1033.1985.tb08986.x · 3.58 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In this paper we report that while 55% of the total post-proline dipeptidyl-aminopeptidase activity in guinea-pig brain is associated with the soluble fraction of the cells, the remaining activity is widely distributed throughout the particulate fractions. A significant portion of this particulate activity is, however, associated with a synaptosomal membrane fraction. The specific activity of this enzyme rose as the synaptosomal membrane fraction was prepared from a synaptosomal fraction and had previously risen at the synaptosomal fraction was prepared from a postmitochondrial pellet. The synaptosomal membrane post-proline dipeptidyl-aminopeptidase was released from the membrane by treatment with Triton X-100 and partially purified by chromatography on Sephadex G-200. By contrast with the soluble enzyme the partially purified solubilised synaptosomal membrane post-proline dipeptidyl-aminopeptidase was not inhibited by 1.0 mM p-chloromercuribenzoate, 1.0 mM N-ethylmaleimide or 0.5 mM puromycin but was inhibited by 0.5 mM bacitracin. The partially purified solubilised enzyme was capable of releasing His-Pro from His-Pro-Val, His-Pro-Leu, His-Pro-Phe and His-Pro-Tyr and of releasing Gly-Pro from Gly-Pro-Ala but could not release Arg-Pro from Arg-Pro-Pro or from Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg (bradykinin). It was also unable to release Pro-Pro from Pro-Pro-Gly or Glp-Pro from Glp-Pro-Ser-Lys-Asp-Ala-Phe-Ile-Gly-Leu-MetNH2 (eledoisin). Using [Pro-3H]thyroliberin we show that the membrane-bound enzyme converts His-ProNH2, produced by the action of the synaptosomal membrane pyroglutamate aminopeptidase, to His-Pro thus competing with the spontaneous cyclisation of His-ProNH2 to His-Pro diketopiperazine. Purified preparations of synaptosomal membrane pyroglutamate aminopeptidase were used to generate His-ProNH2, which could then be converted to His-Pro by the presence of the partially purified synaptosomal membrane post-proline dipeptidyl-aminopeptidase. This preparation was free of contaminating post-proline cleaving endopeptidase, carboxypeptidase P, aminopeptidase P, prolyl carboxypeptidase or proline dipeptidase.
European Journal of Biochemistry 02/1986; 154(2):329-35. DOI:10.1111/j.1432-1033.1986.tb09401.x · 3.58 Impact Factor
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