Identification of [hydroxyproline3]-lysyl-bradykinin released from human plasma protein by kallikrein

[Hydroxyproline3]-lysyl-bradykinin [( Hyp3]-Lys-BK), a new kinin was isolated, besides lysyl-bradykinin (Lys-BK), from the reaction mixture of human plasma protein Cohn's fraction IV-4 with hog pancreatic kallikrein. The liberated kinins were isolated by procedures including ethanol extraction, Sephadex G-15, CM cellulose and reverse-phase high performance liquid chromatography (HPLC) and quantitated by radioimmunoassay. On HPLC, two peaks of immunoreactive kinins emerged. Peak 1, an unknown kinin proceeded to Peak 2 which had an identical retention time to that of Lys-BK. The amino acid sequence of the unknown Peak 1 proved to be Lys-Arg-Pro-Hyp-Gly-Phe-Ser-Pro-Phe-Arg, or [Hyp3]-Lys-BK, and Peak 2 Lys-BK. The ratio of the amounts of two kinins thus formed were [Hyp3]-Lys-BK 25 +/- 4% and Lys-BK 75 +/- 4%. The existence of [Hyp3]-Lys-BK suggests a presence of a new kininogen, containing [Hyp3]-Lys-BK in human plasma protein, possibly undergone post-translational modifications.