Polypeptide components of oligomeric legumin-like thiamin-binding protein from buckwheat seeds characterized by partial amino acid sequencing and photoaffinity labeling |
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Authors: | Rapala-Kozik Maria Ostrowska Katarzyna Bednarczyk Katarzyna Dulinski Robert Kozik Andrzej |
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Institution: | (1) Department of Analytical Biochemistry, Faculty of Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland;(2) Department of Organic Chemistry, Faculty of Chemistry, Jagiellonian University, Ingardena 3, 31-045 Kraków, Poland |
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Abstract: | Among thiamin-binding proteins that ubiquitously occur in plant seeds, that of common buckwheat became a model of extensive studies of the chemical mechanism of ligand-protein interaction. In this work, the polypeptide components of buckwheat seed thiamin-binding protein (BSTBP) are identified and characterized. We suggest that BSTBP is probably a fraction of major storage 13 S globulin (legumin), has an average molecular mass of 235 kDa and comprises hexamers of 57-kDa and 38-kDa subunits in variable combinations. Each subunit is a pair of disulfide-linked polypeptide chains, 36 kDa plus 24 kDa and two-times 22 kDa, respectively. The N-terminal sequences of 22-kDa and 24-kDa components show strict homology with those reported for basic subunits of buckwheat legumin. By photoaffinity labeling of BSTBP with 4-azido-2-nitrobenzoylthiamine, it is shown that the 36-kDa chain plays the major role in thiamin binding, but the other chains may also be variably involved. Putative thiamin-binding fragments are identified and sequenced. |
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Keywords: | Thiamin-binding proteins storage globulins legumins photoaffinity labeling buckwheat seeds |
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