[Binding of Kunitz-Northrop inhibitor on some sorbents].
This work is devoted to the problem of sorption and desorption of Kunitz--Northrop inhibitor on different sorbents. By passing through Dowex 1.10 column 0.05 M glycine-NaOH buffer, pH 10, two fractions with 100% inhibitor activity were identified, while different admixtures and inert proteins remained resin-bound. In trypsin-Spheron 300, trypsin-agarose and anhydrochymotrypsin-Spheron columns the contamination of sorbents-bound inhibitor complex was eliminated by washing out with 0.1 M NaCl, pH 8.0. The resin-bound inhibitor was released at pH 1. The specific activity of the preparation obtained was shown to increase in 200-240-fold, but in the case of anhydrochymotrypsin-Spheron 300 the inhibitors activity was detected both at pH 8.0 and pH 1.7. In this case the increase in specific activity was only 2 and 68-fold, respectively. The most effective inhibitor-peptide desalting was defined at application of dialysis membranes "Spectra/Por" (MWCo: 3000-5000 USA). While applying PAAG electrophoresis the standard SDS-system were shown to be ineffective. Therefore some modifications of this method were used. Being compared with molecular weight of Contrycal and other known peptides this preparation revealed the presence of protein contamination. Within 1-9 mg the peptides demonstrated a linear dependence in trypsin inhibition. The weight and molar ration of inhibitor: trypsin was found to be 1:1 and 3:1, respectively. It was calculated that IUE of the inhibitor had inhibited 0.73 mg of trypsin, IUE of Contrycal--0.15 of the enzyme, that was 4.6 fold less effective than the separated peptide.
Kolodzeĭskaia
M V
MV
Palladin Institute of Biochemistry of NAS of Ukraine, Kyiv. secretar@biochem.kiev.ua
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D010455:Peptides; D010940:Plant Proteins; D014361:Trypsin Inhibitors / Q000378:metabolism*
C053865:Kunitz-type protease inhibitor, plant; D010455:Peptides; D010940:Plant Proteins; D014361:Trypsin Inhibitors