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United States Department of Agriculture

Agricultural Research Service

Title: Cdna Cloning of a Putative Peanut Trypsin Inhibitor with Homology to Peanut Allergens Ara H 3 and Ara H 4

Authors
item Dodo, Hortense - ALABAMA UNIV
item Viquez, Olga - ALABAMA UNIV
item MALEKI, SOHEILA
item Konan, Koffi - ALABAMA UNIV

Submitted to: Journal of Allergy Clinical Immunology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: May 10, 2004
Publication Date: July 23, 2004
Citation: Dodo, H., Viquez, O., Maleki, S.J., Konan, K. 2004. Cdna cloning of a putative peanut trypsin inhibitor with homology to peanut allergens ara h 3 and ara h 4. Journal of Allergy Clinical Immunology.

Interpretive Summary: Peanut allergans and peanut trypsin inhibitors are seed storage proteins. Peanut allergens are known to trigger allergenic reactions. Peanut trypsin inhibitors play an important role in the plant defense mechanism against insects. They are also known to possess anti-nutritional elements, to inhibit trypsin ( a digestive enzyme), and depress growth in animals. Short DNA fragments were synthesized and used to screen a peanut library. Ultimately, three putative trypsin inhibitors were isolated, purified, and sequenced. Sequence analysis of two of these clones revealed a 96% and 93% similiarity with peanut allergens Ara h 4 and Ara h 3, respectively. In conclusion, putative peanut trypsin inhibitors have been isolated and reveal high similiarity at the nucleotide and amino acid level to peanut allergen Ara h 3 and Ara h 4.

Technical Abstract: Background: Peanut allergens and peanut trypsin inhibitors are seed storage proteins. Peanut allergens are known to trigger allergic reactions with symptoms ranging from cutaneous, gastrointestinal, respiratory, systemic to sometimes death. Peanut trypsin inhibitors are pathogenesis-related (PR) proteins and they play an important role in the plant defense mechanism against insects. They are also known to possess antinutritional elements, to inhibit trypsin digestion, and depress growth in animals. Objectives: 1) To synthesize degenerate oligonucleotides based on conserved regions of published amino acid sequences of Bowam-Birk peanut trypsin inhibitors(BBTI) BII and BIII; 2) To screen a peanut cDNA library using the synthesized 32P labeled oligonucleotides as probes; and 3) To isolate, sequence and analyze at least one resulting positive peanut trypsin inhibitor clone. Methods: Thirty-two degenerate oligonucleotides DNA primers of 24 nucleotides each were synthesized based on the published amino acid sequences of peanut BBTI. Two oligonucleotides were selected as probes to screen a peanut Lambda gt 11 cDNA library. Results: Three putative positive clones were isolated, purified, subcloned and sequenced. Sequence analysis revealed a partial cDNA clone of 643 bp with a start codon and with 96% and 93% homology with peanut allergens Ara h 4 and Ara h 3 cDNA clones, respectively. A trypsin inhibitor assay reveals that Ara h 3 allergen protein has trypsin inhibitory activity. Conclusion: In this study, a putative peanut trypsin inhibitor has been isolated and reveals high homology at the nucleotide and amino acid level to peanut allergen Ara h 3 and Ara h 4. At the protein level, it is demonstrated that peanut allergen Ara h 3 also has trypsin inhibitory activity.

Last Modified: 8/19/2014
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