Investigation of kinin generating enzyme in the cardiovascular system.

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Reports have shown that intracardic kinin release exerts anti-ischemic effect. Our study aimed at purification and charaterizatiuon of the kinin-forming enzyme of the dog heart. The ability of the enzyme to generate angiotensin (Ang) II from Ang I was also examined since we previously tissue kallikrein could form Ang II from Ang I.The enzyme from cardiac homogenates has been isolated by a DEAE-Sepharose, aprotinin affinity column, wheat germ lectin-Sepharose 6MB column chromatography. Kininogenase activity was assessed by the measurement of generated kinins with a kinin radioimmunoassay after samples were incubated with bovine low molecular weight kininogen at 37ﾟC for 1 hr. Ang I converting activity was assessed by quantitation of Ang II formed on HPLC by the incubation of the sample with Ang I at 37ﾟC for 3 hrs. The enzyme was electrophoresed on a 12.5% SDS-PAGE followed by Coomassie Brilliant Blue and PAS staining. The purified enzyme is a glycoprotein with an apparent molecular weight of 60 kDa on SDS-PAGE.Kininogenase activity was approximately 22 mug of bradykinin/hr/mg protein at an optimal pH 8.0. The enzyme also converted Ang I to Ang II with a specific activity of approximately 2 mug of Ang II/hr/mg protein at an optimal pH 6.5. Both activity was inhibited by kallikrein inhibitors such as aprotinin, nafamostat, but not by chymostatin and pepstatin.In conclusion, the present study has shown that the kinin-forming enzyme in the dog heart is also able to convert Ang I to Ang II.It is kallikrein-like enzyme different from cathepsin D,cathepsin G,and chymase. This enzyme may play a role in regulating myocardial perfusion through generating kinins and Ang II.Cloning of this enzyme and its localization awaits further examination.