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Irreversible photolabeling of active site of neutral endopeptidase-24. 11 enkephalinase by azidothiorphan and (/sup 14/C)-azidothiorphan

Journal Article · · Mol. Pharmacol.; (United States)
OSTI ID:5520797
; ; ; ;
Azidothiorphan and its (/sup 14/C)-labeled analogue have been developed as photoaffinity ligands for the active site of the neutral endopeptidase 24.11. In in vitro assays azidothiorphan inhibits the endopeptidase activity with a Ki of 0.75 nM. After ultraviolet irradiation the inhibitor binds irreversibly to the enzyme, and many factors suggest that the photolabeling occurs at the active site. The binding is accompanied by a loss of enzymatic activity, and the inclusion of the competitive inhibitor thiorphan protects the endopeptidase from this inactivation. In addition the binding of another competitive inhibitor (/sup 3/H)N-((R,S)-3-hydroxyaminocarbonyl-2-benzyl-1-oxopropyl)-glycine to the active site of endopeptidase-24.11 is inhibited after irradiation with azidothiorphan. Experiments with (/sup 14/C)-azidothiorphan have shown that very little nonspecific binding of inhibitor to enzyme occurs and the the labeled probe remains bound under denaturing conditions. Azidothiorphan has also been found to produce a long-lasting naloxone-reversible analgesia after intracerebroventricular administration. The results show that azidothiorphan should prove useful both for structural studies and for investigations on the synthesis and turnover of the neutral endopeptidase-24.11.
Research Organization:
UA 498 CNRS, Paris, France
OSTI ID:
5520797
Journal Information:
Mol. Pharmacol.; (United States), Journal Name: Mol. Pharmacol.; (United States) Vol. 32:5; ISSN MOPMA
Country of Publication:
United States
Language:
English

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