T‐kinin release from T‐kininogen by rat‐submaxillary‐gland endopeptidase K

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Abstract
Submaxillary gland extracts have been fractionated to characterize the enzyme responsible for the T‐kininogenase activity previously reported in this tissue [Damas, J. & Adam, A. (1985) Mol. Physiol 8, 307–316] and to know whether this activity could be of physiological relevance, since no enzyme reacting in catalytic amounts has been described so far to be able to release a vasoactive peptide from T‐kininogen.The purified enzyme, provisionally called endopeptidase K, has an apparent Mr of 27000 when not reduced prior to analysis but 21000 after reduction and an acidic pI of 4.3 ± 0.1. Antigenically, it is not related to tissue kallikrein. Upon incubation with purified T‐kininogen it may induce a complete liberation of T‐kinin from the precursor provided it is added in stoichiometric amounts. However, in parallel with the liberation of immunoreactive kinin, a proteolysis of T‐kininogen is observed which is not restricted to the site of insertion of T‐kinin as would be expected using a specific kininogenase.In agreement with these results, no change of the mean blood pressure was observed upon injection of endopeptidase K into the circulation of normal rats even if the amount of injected enzyme was up to ten times that required for tissue kallikrein to induce a significant fall in blood pressure.However, in spite of the large proteolysis induced by incubation with stoichiometric amounts of endopeptidase K, the total papain inhibiting capacity of T‐kininogen as well as the value of the apparent inhibition constant, Ki, with this proteinase remained unchanged. Proteolytic fragments which retain cysteine‐proteinase‐inhibiting activity may therefore be released from T‐kininogen by endopeptidase K more easily than immunoreactive kinin, thus emphasizing a prominent function of proteinase inhibitor or of proteinase inhibitor precursor for this molecule.