Presence of a membrane bound pyroglutamyl amino peptidase degrading thyrotropin releasing hormone in rat brain.
In the present work we studied the pattern of degradation of [3H-Pro]-TRH by soluble and membrane fractions from rat brain. Demonstration of the membrane bound or soluble nature of the activities was obtained by comparing their distribution to that of lactate dehydrogenase and by looking at the effect of NaCl washes on the membrane fractions. We observed that the pyroglutamyl amino peptidase activity detected in brain homogenates is a result of two different enzymes. One of them is a soluble enzyme previously characterized, that needs DTT and EDTA for its expression, is inhibited by SH-blocking agents such as iodoacetamide and utilizes p-glu-beta-naphtylamide as a substrate. The other one, a membrane enzyme, is inhibited by chelating agents such as EDTA and DTT, is not affected by iodoacetamide and does not degrade p-glu-beta-naphtylamide. The later presents some specificity towards TRH as shown by competition experiments with TRH analogs. We were able to corroborate that the post proline cleaving enzyme acting on TRH is a soluble enzyme. In membranes we demonstrated also the presence of a post-proline dipeptidyl aminopeptidase. The membrane bound pyroglutamidase activity is a potential new source of L-his-L-pro-diketopiperazine in brain. The presence of a TRH degrading enzyme in membrane fractions is of particular importance in searching an inactivation mechanism of this peptide once it is released into the synaptic cleft.
Garat
B
B
Miranda
J
J
Charli
J L
JL
Joseph-Bravo
P
P
D000626:Aminopeptidases / Q000378:metabolism*; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D001921:Brain / Q000201:enzymology*; D004229:Dithiothreitol / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D066298:In Vitro Techniques; D008297:Male; D008566:Membranes / Q000201:enzymology; D011751:Pyroglutamyl-Peptidase I / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013973:Thyrotropin-Releasing Hormone / Q000097:blood / Q000378:metabolism*
D013973:Thyrotropin-Releasing Hormone; D004492:Edetic Acid; D000626:Aminopeptidases; D011751:Pyroglutamyl-Peptidase I; D004229:Dithiothreitol