Abstract
Background:
Ara h 2 is a major peanut allergen recognized by IgE in more than 90% of patients. After electrophoretic separation the purified protein exists as a doublet, and sequences of one incomplete cDNA and one genomic clone for this allergen have been reported.
Methods:
Ara h 2 isoforms were purified and analyzed by mass spectroscopy, and PCR amplification products of Ara h 2 were cloned and sequenced.
Results:
Mass spectroscopy of purified Ara h 2 clearly identified a molecular doublet of 16,670 and 18,050 Daltons. Amplification of a peanut cDNA library using PCR primer pairs located at the amino- and carboxy-terminus revealed 2 bands separated by 50 base pairs, which we cloned and sequenced. Two types of complete cDNA clones were obtained, Ara h 2.01 and Ara h 2.02. Compared to Ara h 2.01 and the previously reported cDNA sequences, Ara h 2.02 is characterized by a 12 amino acid insertion starting at position 75 that contains a third repeat of the major IgE binding epitope DPYSPS.
Conclusion:
We demonstrated the molecular and genetic characteristics of two Ara h 2 isoforms, revealing that one, Ara h 2.02, might be the more potent allergen.
Copyright 2003 S. Karger AG, Basel
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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2S Albumins, Plant
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Amino Acid Sequence
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Antigens, Plant
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Cloning, Molecular
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Codon, Initiator / classification
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Codon, Initiator / genetics
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Codon, Initiator / isolation & purification
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Codon, Terminator / classification
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Codon, Terminator / genetics
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Codon, Terminator / isolation & purification
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DNA Primers / classification
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DNA Primers / genetics
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DNA Primers / isolation & purification
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DNA Transposable Elements / genetics
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DNA Transposable Elements / immunology
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DNA, Complementary / classification*
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DNA, Complementary / genetics
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DNA, Complementary / isolation & purification*
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Glycoproteins / classification*
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Glycoproteins / genetics
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Glycoproteins / isolation & purification*
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Humans
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Immunoglobulin E / genetics
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Immunoglobulin E / immunology
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Mass Spectrometry
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Molecular Sequence Data
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Molecular Weight
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Open Reading Frames / genetics
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Plant Proteins
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Polymerase Chain Reaction
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Protein Isoforms / classification
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Protein Isoforms / genetics
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Protein Isoforms / isolation & purification
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Terminal Repeat Sequences / genetics
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Terminal Repeat Sequences / immunology
Substances
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2S Albumins, Plant
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Antigens, Plant
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Ara h II protein, Arachis hypogaea
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Codon, Initiator
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Codon, Terminator
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DNA Primers
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DNA Transposable Elements
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DNA, Complementary
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Glycoproteins
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Plant Proteins
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Protein Isoforms
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Immunoglobulin E