| Project/Area Number |
13660012
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
作物学
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| Research Institution | HOKKAIDO UNIVERSITY |
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Principal Investigator |
FUJINO Kaien Hokkaido Univ., Grad. School of Agr., Lecturer, 大学院・農学研究科, 講師 (80229020)
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| Co-Investigator(Kenkyū-buntansha) |
MASUDA Kiyoshi Hokkaido Univ., Grad. School of Agr., Asso. Prof., 大学院・農学研究科, 助教授 (60157203)
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| Project Period (FY) |
2001 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2002: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 2001: ¥2,400,000 (Direct Cost: ¥2,400,000)
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| Keywords | buckwheat / legumin / allergen / seed storage protein / 2S albumin / 種子タンパク質 / そばアレルギー |
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| Research Abstract |
Buckwheat (Fagopyrum esculenturn Moench) genes FA02 and FA18 were isolated from immature seeds harvested 14 days after polination, and were found to encode legumin-like proteins that are expressed during seed development. The deduced amino acid sequence of FA02 was identical to the N-terminal amino acid domain of BW24KD, which is believed to be a major buckwheat allergen. It was predicted that FA02 would be cleaved to generate two separate components, a 41.3 kDa alpha-subunit and a 21 kDa beta-subunit. It was deduced from molecular mass and N terminal amino acid that BW24KD was the beta-submit of FA02. Antiserum was raised against the deduced FA02 beta-subunit and immunoblotting of total protein from buckwheat seeds (Fagopyrum esculentum M. and Fagopyrum tartaricum M.) revealed that several groups of proteins reacted with the antiserum. Based on the N-terminal amino acid sequence of the 10 kDa buckwheat allergenic protein (BW10KD) which strongly reacted with IgE in 50% of allergic patients, a degenerate primer were synthesized and 3'RACE PCR amplification was carried out. Fe2SA1 cDNA clone, which was isolated from a cDNA library made of immature buckwheat seeds using as a probe a3' RACE PCR clone, was partially identical to 2S albumin seed storage protein. The deduced amino acid sequence of Fe2SA1 was partially identical to the 16 and 18 kDa molecule, which are believed to be major buckwheat allergens. In an E. coli. Expression system, the translation product of a cDNA encoding the Fe2SA1 was strongly recognized by sera from allergic patients.
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