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Effect of cysteine mutagenesis on human IgE reactivity of recombinant forms of the major rye grass pollen allergen Lol p 1

Authors

  • Nicole De Weerd,

    1. Plant Molecular Biology and Biotechnology Laboratory, Institute of Land and Food Resources, University of Melbourne, Parkville, Victoria, Australia
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  • Prem L Bhalla,

    1. Plant Molecular Biology and Biotechnology Laboratory, Institute of Land and Food Resources, University of Melbourne, Parkville, Victoria, Australia
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  • Mohan B Singh

    1. Plant Molecular Biology and Biotechnology Laboratory, Institute of Land and Food Resources, University of Melbourne, Parkville, Victoria, Australia
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Associate Professor Mohan B Singh, Plant Molecular Biology and Biotechnology Laboratory, Institute of Land and Food Resources, The University of Melbourne, Victoria 3010, Australia. Email: m.singh@landfood.unimelb.edu.au

Abstract

Background:  Hypersensitivity to the group 1 grass pollen allergens is a significant causative factor in the onset of symptoms for hay fever sufferers. To better understand the IgE reactivity of the group 1 allergen from rye grass pollen, we sought to disrupt potential conformational IgE epitopes on recombinant (r) Lol p 1 by the specific replacement of the seven cysteine residues in the protein sequence.

Methods:  Site-directed mutagenesis on the Lol p 1 coding sequence was used to replace all seven cysteine residues with serine residues. rLol p 1 and the seven cysteine variants generated by this method were tested for comparative human IgE reactivity via western blot immunoscreening and densitometry.

Results:  Alteration of the cysteine residues at amino acid positions 72, 77, 83 and 139 of rLol p 1 was found to reduce the human IgE binding potential of the molecule. However, the most consistent reduction in human IgE reactivity was demonstrated by replacement of C77; human IgE antibodies showed an average 62.7% reduction in reactivity to this molecule.

Conclusions:  The present investigation has shown that at least one of the cysteine residues within the Lol p 1 protein contributes to the IgE binding properties of this allergen.

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