Molecular cloning, recombinant expression, and immunological characterization of a novel allergen from tartary buckwheat.
Buckwheat is generally regarded as a nutritionally rich food source. However, earlier studies prove that it also causes allergies to subjects. Allergenic proteins with a strong IgE-binding activity have been identified in common buckwheat (CB) and a 24 kDa allergen (rTBa) in tartary buckwheat (TB). The objective of this research was to clone and express a novel allergen in tartary buckwheat and to evaluate its structure and immunological activity. The 1773 bp full-length cDNA was amplified and cloned from the total RNA of TB by polymerase chain reaction (PCR) and rapid amplification of cDNA ends (RACE) methods. Its nucleotide sequence had high similarity with legume-like 13S storage protein mRNA in CB. The deduced amino acid sequence included a putative signal peptide and 18 fragments as its epitope sites. The predicted full-length TB allergen sequence was found to have two domains, and the recombinant protein reacted with sera from patients with positive IgE binding to buckwheat and had a lower binding ability than the recombinant TBa and recombinant TBb (C- and N-terminal amino acid sequence of TBt codes for protein). This fact suggests that full-length TB allergen may hydrolyze to two domains in vivo, decreasing the IgE-binding ability.
Zhang
Xin
X
Key Laboratory for Chemical Biology and Molecular Engineering of the Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan, China.
Yuan
Jing Ming
JM
Cui
Xiao Dong
XD
Wang
Zhuan Hua
ZH
D000595:Amino Acid Sequence; D052179:Antigens, Plant / Q000235:genetics* / Q000276:immunology*; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry; D018744:DNA, Plant / Q000235:genetics; D019612:Fagopyrum / Q000737:chemistry* / Q000276:immunology; D005512:Food Hypersensitivity / Q000276:immunology; D015870:Gene Expression; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000235:genetics*; D017422:Sequence Analysis, DNA
D052179:Antigens, Plant; D018076:DNA, Complementary; D018744:DNA, Plant; D010940:Plant Proteins; D011994:Recombinant Proteins; D007073:Immunoglobulin E