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PURIFICATION AND PARTIAL CHARACTERIZATION OF TWO PROTEINACEOUS α‐AMYLASE INHIBITORS FROM TRITICALE
Author(s) -
IDA ELZA I.,
FINARDIFILHO FLAVIO,
LAJOLO FRANCO M.
Publication year - 1994
Publication title -
journal of food biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.507
H-Index - 47
eISSN - 1745-4514
pISSN - 0145-8884
DOI - 10.1111/j.1745-4514.1994.tb00491.x
Subject(s) - amylase , chemistry , aspergillus oryzae , chromatography , triticale , chymotrypsin , pepsin , biochemistry , isoelectric point , sephadex , trypsin , isoelectric focusing , enzyme , biology , botany
A total of six α‐amylase inhibitory proteins (isoinhibitors) were extracted from triticale (Triticum X Secale) seeds and two of them were purified to homogeneity. The isoinhibitors were extracted by 70% ethanol and produced, by Sephadex G‐100 chromatography, two peaks that exhibited α‐amylase inhibitory activity. Further purification of the most active peak by DEAE‐cellulose chromatography resulted in six active fractions. Two of them designated as TAI‐5 and TAI‐6, considered to be homogeneous by both acidic and alkaline electrophoresis, were partially characterized. The isoelectric points were 4.80 and 4.70, and the molecular weights 39, 200 and 29, 200, respectively. Under dissociating conditions the molecular weights were 13, 500 and 13, 000, suggesting that the isoinhibitors are composed of three and two subunits, respectively. Both isoinhibitors were stable at different pHs, relatively stable at 98C, and resistant to proteolysis by trypsin, chymotrypsin and pepsin. The optimum interaction pH for both isoinhibitors with human salivary amylase was 7.9. They exhibited specificity to human salivary and porcine pancreatic α‐amylases, but had no inhibitory activity on Bacillus subtillis, Aspergillus oryzae and endogenous triticale α‐amylases.