Cereal Chem 48:303 - 311.  |  VIEW ARTICLE
Isoelectric Point Differences in Commercial Soybean Trypsin Inhibitors.

L. C. Wang. Copyright 1971 by the American Association of Cereal Chemists, Inc. 

Isoelectric points of Kunitz' crystalline soybean trypsin inhibitor (STI), Bowman-Birk STI, and STI prepared by chromatography on diethylaminoethyl (DEAE)-cellulose were investigated by isoelectric focusing, gel filtration, ion-exchange resin treatment, solubility test, and titration. Results suggest that the STI of Kunitz may have an isoelectric point below pH 4.5. The isoelectric focusing pattern of the Kunitz inhibitor showed three peaks: at pH 3.5, 3.7, and 4.4. All three proteins were active inhibitors. Chromatography on anion-exchange resins separated the Kunitz STI into two fractions: one that eluted with water and focused at pH 3.5 and another that eluted with increasing concentrations of NaCl solution and focused at pH 4.0. When ion-retardation resins were used only one peak resulted, focusing at pH 3.8. Solubility tests on the Kunitz preparation before and after treatment with anion-exchange resins supported evidence for different isoelectric points. Titration of Kunitz' STI at three levels of ionic strength gave an isoionic zone lower than pH 4.5. The Kunitz inhibitor contained about 20% impurities based on gel filtration and the purified protein focused at a single isoelectric pH of 4.0. The Bowman-Birk inhibitor consistently showed an isoelectric point of pH 4.2 in focusing and solubility tests, and it had little or not impurities on gel filtration. Although STI prepared by chromatography on DEAE-cellulose contained impurities, heterogeneity of isoelectric point (pH 4.0) was less than for Kunitz' STI. The multiple isoelectric points of crystalline STI were believed caused by interaction between protein and other constituents, possibly contaminants.