Chicken liver Pz-peptidase, a thiol-dependent metallo-endopeptidase

Abstract

Pz-peptidase was purified from chicken liver as a protein of Mr 80000 and pl 5.2. The purified enzyme hydrolysed phenylazobenzyloxycarbonyl-Pro-Leu-Gly-Pro-D-Arg, 2,4-dinitrophenyl-Pro-Leu-Gly-Pro-Trp-D-Lys, 7-methoxycoumarin-3-carboxylyl-Pro-Leu-Gly-Pro-D-(2,4-dinitrophenyl)Lys, benzoyl-Gly-Ala-Ala-Phe-p-aminobenzoate, Ac-Ala4 (at the Ala-I-Ala-2 bond) and bradykinin (at the Phe-5-Ser-6 bond). No hydrolysis of proteins was detected. Loss of activity in the presence of EDTA or 1,10-phenanthroline was time-dependent. Metal ions found to restore activity after treatment with EDTA were Zn2+, Mn2+, Ca2+, Co2+ and Cd2+, in decreasing order of effectiveness. Ni2+, Fe2+ and higher concentrations of Zn2+ were inhibitory. Inhibition by N-[1-(RS)-carboxy-3-phenylpropyl]-Ala-Ala-Tyr-p-aminobenzoate and related compounds showed K1 values (down to 5 nM) somewhat lower than those for the rat enzyme. Pz-peptidase was activated by low concentrations of 2-mercaptoethanol and dithiothreitol, but inhibited by higher concentrations. pChloromercuribenzoate and some other thiol-blocking reagents were inhibitory. Inactivation by diethyl pyrocarbonate that was reversible by hydroxylamine showed the presence of essential histidine residue(s). We conclude that chicken Pzpeptidase is a metallo-endopeptidase with thiol-dependence. Moreover, the properties of chicken Pz-peptidase agree well with those described for mammalian soluble metallo-endopeptidase and endo-oligopeptidase A, consistent with the view that these three types of activity are all attributable to the single enzyme for which the niame thimet peptidase has been proposed.

Mammalian microsomal and soluble Ras‐processing peptidase activities are distinct

“…When the preincubation time with o-phenanthroline was increased to 60 min at room temperature, the IC50 decreased from 300 ~tM to < 100 ~tM. In other systems, inhibition by EDTA has also been reported to be time dependent [25], although in the present study this chelator did not inhibit soluble peptidase activity up to 1 raM. It is noted that the metalloprotease carboxypeptidase B, though sensitive to ophenanthroline, is also resistant to EDTA [26].…”

Purification of the main somatostatin‐degrading proteases from rat and pig brains, their action on other neuropeptides, and their identification as endopeptidases 24.15 and 24.16

“…The acronym 'thimct oligopeptidase' (thiol-and metal-dependent oligopeptidase) has been introduced for these enzymes [26,291. However, the identity of endo-oligopeptidase A (or Pz-peptidase) with endopeptidase 24.15 has been opposed [30,31].…”

Two Thimet Oligopeptidase-Like Pz Peptidases Produced by a Collagen- Degrading Thermophile, Geobacillus collagenovorans MO-1

“…In spite of the fact that the monomer of tripeptide [Gly-Pro-Leu] was not hydrolyzed, the dimer [Gly-Pro-Leu] 2 , trimer [Gly-Pro-Leu] 3 , and tetramer [Gly-ProLeu] 4 peptides were hydrolyzed by both Pz peptidases between Leu-Gly, suggesting that the enzymes should have a significant role in collagen degradation. The pentamer [Gly-Pro-Leu] 5 and longer peptides were too insoluble in water to perform the assay. From these results, Pz peptidases A and B are endopeptidases and fail to hydrolyze less than tetrapeptides.…”