title: Mechanism of ligand-protein interaction in plant seed thiamine-binding proteins. Preliminary chemical identification of amino acid residues essential for thiamine binding to the buckwheat-seed protein.
abstract: Thiamine-binding protein, isolated from buckwheat seeds, was chemically modified in an attempt to identify amino acid residues involved in protein-thiamine interaction. No evidence was found in support of specific roles of arginine residues, sulfhydryl groups, amino groups and tyrosine residues. Under carefully controlled reaction conditions (Tris pH 5-6), the modification with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide caused a complete loss of thiamine-binding capacity. Thus, the carboxyl groups seemed to be essential for binding, possibly for ionic interaction with protein-bound thiamine cation. A selective modification of histidine residues using diethylpyrocarbonate correlated with a loss of thiamine-binding capacity; the modification and the loss of binding capacity could be reversed with hydroxylamine; some ligand-protection against modification was observed. From Tsou analysis of diethylpyrocarbonate modification and resulting loss of thiamine-binding it was suggested that 1-2 of 20 histidine residues of the protein were essential for thiamine binding. The essential histidine(s) might be present in the binding site and possibly were involved in hydrogen bonding(s) with protein-bound thiamine molecule.
authors.affiliation: Jan Zurzycki Institute of Molecular Biology, Jagiellonian University, Kraków, Poland.
authors.forename: M
authors.initials: M
authors.lastname: Rapala-Kozik
authors.forename: A
authors.initials: A
authors.lastname: Kozik
mesh_terms: D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism; D006639:Histidine; D010940:Plant Proteins / Q000378:metabolism*; D012639:Seeds; D013329:Structure-Activity Relationship; D013831:Thiamine / Q000378:metabolism*; D014908:Triticum; D014443:Tyrosine
chemical_list: D002352:Carrier Proteins; D010940:Plant Proteins; C026989:thiamine-binding protein; D014443:Tyrosine; D006639:Histidine; D013831:Thiamine