Epitope mapping and immunological characterization of a major allergen TBa in tartary buckwheat.
Predicted by an antigenic program, full-length tartary buckwheat allergen (TBa) is divided into six fragments: E1, E2, E12, E3, E4 and E34. Immunological assays revealed that E1 has the greatest binding activity to patients' serum IgE. Five mutants of E1 gene (L39R, L42R, L47R, V52R and L54R) were constructed using site-directed mutagenesis and each protein was expressed in Escherichia coli BL21. Following purification by Ni(2+) affinity chromatography, ELISA and dot-blot were performed for wild type E1 and its mutants using sera from buckwheat allergic patients and healthy controls. Mutants L42R, L47R, and L54R had weaker IgE binding activity to patient's sera than wild-type E1 implying that Leu42, Leu47, and Leu54 might be involved in the allergic activity of TBa.
Ren
Xiaoxia
X
Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Institute of Biotechnology, Shanxi University, 030006 Taiyuan, People's Republic of China.
Zhang
Xin
X
Li
Yuying
Y
Wang
Zhuanhua
Z
D000485:Allergens / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D004926:Escherichia coli / Q000235:genetics; D019612:Fagopyrum / Q000737:chemistry*; D015870:Gene Expression; D006801:Humans; D015151:Immunoblotting; D007073:Immunoglobulin E / Q000097:blood / Q000276:immunology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010940:Plant Proteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D011485:Protein Binding; D016415:Sequence Alignment
D000485:Allergens; D010940:Plant Proteins; D007073:Immunoglobulin E