Activation mechanism of human urinary prokallikrein using trypsin as a model activator

Rapid release of a small peptide from human urinary prokallikrein by trypsin resulted in activation of the prokallikrein. The peptide was identified as the propeptide of the kallikrein from its amino acid sequence. Two large disulfide-linked peptides were also produced very slowly, which accompanied the increase in kallikrein activity. The molecular weights of the two peptides were roughly estimated to be 18,000 and 25,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). N-Terminal amino acid sequences were determined as Ile-Val-Gly-Gly-Trp-Glu-Cys-Glu-Gln-His for the Mr 18,000 peptide and Gln-Ala-Asp-Glu-Asp-Tyr-Ser-His-Asp-Leu for the Mr 25,000 peptide. The N-terminal sequence of the Mr 18,000 peptide was identical to that of the kallikrein. Both peptides contained carbohydrate side chains as judged by staining with periodic acid-Schiff's base. The results indicate strongly that trypsin hydrolyses two specific bonds of human urinary prokallikrein selectively, which are cleaved upon physiological activation to yield the two-chain kallikrein.