Pharmacological Activities and Hydrolysis by Peptidases of [Phospho-Ser6]-Bradykinin (pS6-BK)☆

Abstract Phosphorylated kininogen and some of its fragments containing serine phosphorylated bradykinin ([pS 6 ]-Bk) were identified in human serum and plasma by a phosphoproteomic approach. We report the kininogenase ability of human tissue and plasma kallikreins and tryptase to generate [pS 6 ]-Bk or Lys-[pS 6 ]-Bk having as substrate the synthetic human kininogen fluorescent fragment Abz-MISLMKRPPGF[pS 386 ]PFRSSRI-NH2. The pharmacological assays of [pS 6 ]-Bk showed it as a full B2 bradykinin receptor agonist in smooth muscle, it produces a portal liver hypertensive response in rat and mouse paw edema that lasts longer than Bk. The rat hypotensive response to infusions of Bk is greater than that of [pS 6 ]Bk, both if injected through femoral vein or aorta. [pS 6 ]-Bk was more resistant than Bk to kininase digestion performed with angiotensin converting enzyme, neprilysin, thimet oligopeptidase, aminopeptidase P and carboxypeptidase M. 1 H-NMR experiments indicated that [pS 6 ]-Bk has lower flexibility, with the pS 6 -P 7 bond restricted to the trans conformation, and can explain [pS 6 ]-Bk resistance to hydrolysis. In conclusion, [pS 6 ]-Bk presenting lower activity than Bk, with longer lasting effects and being slowly released by kininogenases from synthetic Abz-MISLMKRPPGF[pS 386 ]PFRSSRI-NH 2 , suggests that phosphorylation of the kininogens can be an efficient kallikrein-kinin system regulator.