Purification and characterization of tonin.
Tonin was purified from rat submaxillary glands by differential centrifugation, ammonium sulphate precipitation, gel filtration on Sephadex G150, and by ion-exchange chromatography on DEAE-cellulose, phospho-cellulose, SP-Sephadex C25, and SP-Sephadex C50. Purified tonin was shown to be homogeneous by analytical electrophoresis and by analytical ultracentrifugation analysis. Purified tonin was very stable when stored in buffers of low pH values or when incubated at high temperatures in neutral solution. The molecular weight estimated by sedimentation equilibrium was 28 700. The pH optimum was near 6.8 in a 0.1 M potassium phosphate buffer. The Michaelis-Menten constant for tonin using angiotensin I as substrate was about 4 X 10(-5) M. Tonin activity was strongly inhibited by plasma. Kinetic studies using angiotensin I as substrate showed that the inhibition of tonin by plasma was of the non-competitive type.
Purification and characterization of tonin. Tonin was purified from rat submaxillary glands by differential centrifugation, ammonium sulphate precipitation, gel filtration on Sephadex G150, and by ion-exchange chromatography on DEAE-cellulose, phospho-cellulose, SP-Sephadex C25, and SP-Sephadex C50. Purified tonin was shown to be homogeneous by analytical electrophoresis and by analytical ultracentrifugation analysis. Purified tonin was very stable when stored in buffers of low pH values or when incubated at high temperatures in neutral solution. The molecular weight estimated by sedimentation equilibrium was 28 700. The pH optimum was near 6.8 in a 0.1 M potassium phosphate buffer. The Michaelis-Menten constant for tonin using angiotensin I as substrate was about 4 X 10(-5) M. Tonin activity was strongly inhibited by plasma. Kinetic studies using angiotensin I as substrate showed that the inhibition of tonin by plasma was of the non-competitive type.