Molecular and immunochemical evidences demonstrate that endooligopeptidaseA is the predominant cytosolic oligopeptidase of rabbit brain

Oligopeptidases are tissue endopeptidases that do not attack proteins and a re likely to be involved in the maturation and degradation of peptide hormo nes and neuropeptides. The rabbit brain endooligopeptidase A and the rat te stes soluble metallopeptidase (EC 3.4.24.15) are thiol-activated oligopepti dases which are able to generate enkephalin from a number of opiod peptides and to inactivate bradykinin and neurotensin by hydrolyzing the same pepti de bonds. A monospecific antibody raised against the purified rabbit brain endooligopeptidase A allowed the identification of a 2.3 kb cDNA coding for a truncated enzyme of 512 amino acids, displaying the same enzymatic featu res as endooligopeptidase A. In spite of all efforts, employing several str ategies, the full-length cDNA could not be cloned until now. The analysis o f the deduced amino acid sequence showed no similarity to the rat testes me talloendopeptidase sequence, except for the presence of the typical metallo protease consensus sequence [HEXXH]. The antibody raised against recombinan t endooligopeptidase A specifically inhibited its own activity and reduced the thiol-activated oligopeptidase activity of rabbit brain cytosol to less than 30%. Analysis of the endooligopeptidase A tissue distribution indicat ed that this enzyme is mainly expressed in the CNS, whereas the soluble met allo EC 3.4.24.15 is mainly expressed in peripheral tissues. (C) 2000 Acade mic Press.