• Applied Biological Chemistry
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• v.33 no.4
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• pp.287-292
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• 1990

Eight(I through VII) of Bowman-Birk proteinase inhibitor have been isolated from soybean with DEAE-Sephadex A-50. Inhibitor VII was a typical BBTI, showing high cysteine content(17%/mole) ud low trypsin to chymotrypsin inhibiting activity(TIA/CIA= 1.0) with the independent reactive site to each enzyme. Dissociation constant of trypsin-BBTI and chymotrypsin-BBTl complexes were $9.17{\times}10^{-9}M$ and $5.14{\times}10^{-8}M$, respectively. Inhibitor Vll was extremely heat stable. Six hours heat treatment at $100^{\circ}C$ caused only 50% decrease in it's original inhibiting activity. Except inhibitor III,6 other isoinhibitors differed from a typical BBTI in TIA/CIA, values, ranging from 3 to 29.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.49 no.4
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• pp.342-348
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• 2004

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the $60\%$ isopropanol extract of soybean(Glycine max [L.] Merr.) seed revealed two abundant proteins with molecular masses of 19 and 10 kDa. Amino acid analysis revealed that the isopropanol-extractable protein fraction was rich in cysteine. Two-dimensional gel electro-phoretic analysis indicated that the 19kDa and 10kDa proteins had pI of 4.2 and 4.0 respectively. Peptide mass fingerprints of trypsin digests of the two proteins obtained using matrix-assisted, laser desorption/ionization-time of flight (MALDI-TOF) mass spectroscopy revealed the 19kDa protein was Kunitz trypsin inhibitor and the 10kDa protein was Bowman-Birk proteinase inhibitor. When resolved under non-denaturing conditions, the isopropanol-extracted proteins inhibited trypsin and chymotrypsin activity. Results presented in this study demonstrate that isopropanol extraction of soybean seed could be used as a simple and rapid method to obtain a protein fraction enriched in Kunitz trypsin and Bowman-Birk proteinase inhibitors. Since proteinase inhibitors are rich in sulfur amino acids and are putative anticarcinogens, this rapid and inexpensive isolation procedure could facilitate efforts in nutrition and cancer research.

• Applied Biological Chemistry
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• v.32 no.3
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• pp.222-231
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• 1989

Three classes of proteinase inhibitors are known in soybean; the Kunitz trypsin inhibitor (SKTI), the Bowman-Birk proteinase inhibitor and its isoinhibitors. To study the molecular structure and expression characteristics of the SKTI, antibody was obtained by immunizing rabbit with the SKTI purified from soybean by preparative electrophoresis. Anti-SKTI antibody was not only specific for mature SKTI in soybean seed but also recognized the precursor which was synthesized in vitro. Translation in vitro was carried out in wheat germ extract with polyadenylated mRNA isolated from developing soybean seeds. One of the seed specific translation products, MW 24K, was identified to be the precursor for the SKTI by immunoprecipitation with anti-SKTI antibody. Mature SKTI of MW 20K, however, was not detected in the translates in vitro. These results suggest that the precursor polypeptide is synthesized from the mRNA and is cleaved to yield mature SKTI in soybean seed. The SKTI gene was expressed with the maturation of soybean seed in a tissue-specific and development stage-specific manner.