ISOLATION AND CHARACTERIZATION OF THE 11S GLOBULIN FROM AMARANTH SEEDS

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Abstract
Globulins of Amaranthus hypochondriacus were extracted with two different buffer systems which varied in ionic strength. SDS-PAGE analysis demonstrated that subunit patterns were different between the two extracting systems. Amarantin, the native 11S globulin of amaranth, and its subunits were purified by gel filtration chromatography and preparative electrophoresis. The native amarantin exhibited two heterogeneous forms of MW 330 and 400 kDa, which yielded the same subunit composition after reduction. SDS-PAGE analysis showed that prior to reduction, the 50–52 kDa subunit was the major band, and that after reduction, two new bands of 32–34 kDa and 22–24 kDa appeared. This is a typical characteristic of 11S proteins whose subunits consist of an acidic polypeptide (27–37 kDa) and a basic polypeptide (20–24 kDa) linked by a disulfide bond. Ultracentrification analysis showed that amarantin has a 11.9S sedimentation coefficient whereas DSC demonstrated that the denaturation temperatures in the presence of H2O, Tris-HCl and K2HPO4-KH2PO4 are 99C, 99.8C and 103C, respectively.