HIGH-MOLECULAR-WEIGHT KININOGEN PEPTIDES INHIBIT THE FORMATION OF KALLIKREIN ON ENDOTHELIAL-CELL SURFACES AND SUBSEQUENT UROKINASE-DEPENDENT PLASMIN FORMATION

A sequence of 31 amino acids (S565-K595) in domain 6 of the light chai n of high molecular weight kininogen (HK) has previously been shown to be responsible for the binding of plasma prekallikrein (PK) or kallik rein, To find effective peptides that might block binding between HK a nd PK on cell surfaces, a new series of synthetic peptides has now bee n prepared that incorporates portions of this binding domain sequence. For mapping the minimal sequence within HK, these new peptides were t ested for their ability to compete with HK for binding PK in a cell-fr ee system and on human umbilical vein endothelial cells (HUVEC), In th e former, at pH 7.4, the kds for binding between kallikrein and either D567-K595, S565-P594, D567-S593, or D567-T591 were all similar to tha t for the binding of S565-K595 (0.2 to 0.4 mu mol/L), but those for th e binding of D568-K595, W569-K595, and D567-P589 were an order of magn itude greater (kd = 2 to 5 mu mol/L). D567-S586, the shortest chain le ngth of the N-and C-terminal truncation sequences tested, does not eff ectively compete with kininogen for kallikrein binding (kd = 100 mu mo l/L). These results imply that D567-T591, a 25-residue peptide (HK25c) , contains sufficient structural information for binding kallikrein in solution. D567-T591 also is the minimum structural sequence to block binding of kallikrein to HUVEC-bound HK (IC50 = 50 nmol/L) and to inhi bit PK activation to kallikrein on the cell surface (IC50 = 80 nmol/L) . In addition, D567-T591 also inhibits the generation of kal-likrein-a ctivated urokinase, which activates plasminogen to plasmin (IC50 = 100 nmol/L), Thus, HK-derived peptides may be useful compounds for modula ting excessive fibrinolysis and hypotension in sepsis and multiple tra uma. (C) 1997 by The American Society of Hematology.