Purification and partial characteristic of a major gliadin-degrading cysteine endopeptidase from germinating triticale seeds |
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Authors: | Email author" target="_blank">Beata?PrabuckaEmail author Wies?aw?Bielawski |
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Institution: | (1) Department of Biochemistry, Warsaw Agricultural University, Nowoursynowska 159, 02-776 Warszawa, Poland |
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Abstract: | The endopeptidase of the highest electrophoretic mobility was the main endopeptidase hydrolyzing gliadin in the endosperm
of germinated triticale (X Triticosecale Wittm.) grains after three days of imbibition. Activity of this endopeptidase, named EP8 starts to be detectable after two
days of imbibition. The appearance of its activity in the endosperm on a second day of imbibition may suggest that EP8 is
synthesized in aleurone during germination and/or secreted into the starchy endosperm as an inactive polypeptide during grains
development and then activated. EP8 was isolated from the endosperm of germinating triticale seeds and purified 257-fold using
ammonium sulphate, ion exchange chromatography on DEAE Sepharose CL-6B and gel filtration on Sephadex G-100. The enzyme was
totally inhibited by E-64—class-specific cysteine proteinases inhibitor and activated by thiol compounds. Molecular weight
estimated by SDS-PAGE was 39.5 kDa. The optimum pH for the hydrolysis of gliadin was 4.2 and for hemoglobin 5.2. High activity
of EP8 against wheat gliadin in vitro suggests that this cysteine endopeptidase plays a major role in the mobilization of storage proteins in the endosperm of
germinating triticale grains. |
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Keywords: | cereals cysteine endopeptidases germination endopeptidase purification triticale |
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