Degradation of cholecystokinin octapeptide by the neutral endopeptidase EC 3.4.24.11 and design of proteolysis-resistant analogues of the peptide.
Inactivation of cholecystokinin octapeptide in vitro involves a metalloendopeptidase (EC 3.4.24.11) also called enkephalinase that inactivated the peptide both by a sequential pathway of hydrolysis (removal of Phe-NH(2) followed by cleavage of Trp-Met-Asp) and by an endopeptidase action (production of the tetrapeptides). As enkephalinase cleaved CCK-8 at the Gly(4)-Trp(5), Trp(5)-Met(6) and Asp(7)-Phe(8) bonds, we investigated the stability of analogues having: (1) substitutions of l amino acids by a d stereoisomer, (2) a substitution of Asp(7) by a ? Ala residue and (3) modifications of the Trp residue obtained by replacing the nitrogen atom in the indol ring by either an oxygen ([Bfa(5)]CCK-8) or a sulphur atom ([Bta(5)]CCK-8). Among these different CCK derivatives, [?Ala(7)], [dMet(6)] and [dTrp(5)]CCK-8 were not hydrolyzed by enkephalinase: [dAla(d)]CCK-8 was rapidly cleaved by the enzyme. [Bta(5)] and [Bfa(5)]CCK-8 did not prove to be quite resistant; however the C-terminal tetrapeptides having the same modifications on the Trp residue were not cleaved although they interacted with the enzyme binding site. The stability and biological activity of the peptidase-resistant analogues of CCK-8 remain to be determined in vivo.
Najdovski
T
T
Laboratoire Pluridisciplinaire de Recherche Expérimentale Biomédicale, Faculté de Médecine, Campus Erasme, Brussels University, Brussels, Belgium.
de Pont
J J
JJ
Tesser
G I
GI
Penke
B
B
Martinez
J
J
Deschodt-Lanckman
M
M