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Plant-based Heterologous Expression of Mal d 2, a Thaumatin-like Protein and Allergen of Apple (Malus domestica), and its Characterization as an Antifungal Protein
- Source :
-
Journal of Molecular Biology . Jun2003, Vol. 329 Issue 4, p721. 10p. - Publication Year :
- 2003
-
Abstract
- Mal d 2 is a thaumatin-like protein and important allergen of apple fruits that is associated with IgE-mediated symptoms in apple allergic individuals. We obtained a full-length cDNA clone of Mal d 2 from RNA isolated from ripe apple (Malus domestica cv. Golden Delicious). The cDNA''s open reading frame encodes a protein of 246 amino acid residues including a signal peptide of 24 residues and two putative glycosylation sites. The deduced amino acid sequence of the mature Mal d 2 protein results in a predicted molecular mass of 23,210.9 Da and a calculated pI of 4.55. Sequence comparisons and molecular modeling place Mal d 2 among those pathogenesis-related thaumatin-like proteins that contain a conserved acidic cleft. In order to ensure the correct formation of the protein''s eight conserved disulfide bridges we expressed Mal d 2 in Nicotiana benthamiana plants by the use of a tobacco mosaic viral vector. Transfected N. benthamiana plants accumulated Mal d 2 to levels of at least 2% of total soluble protein. MALDI-TOF mass spectrometric analyses of the recombinant Mal d 2 and its proteolytic fragments showed that the apple-specific leader peptide was correctly cleaved off by the host plant and that the mature recombinant protein was intact and not glycosylated. Purified recombinant Mal d 2 displayed the ability to bind IgE from apple-allergic individuals equivalent to natural Mal d 2. In addition, the recombinant thaumatin-like Mal d 2 exhibited antifungal activity against Fusarium oxysporum and Penicillium expansum, implying a function in plant defense against fungal pathogens. [Copyright &y& Elsevier]
- Subjects :
- *XENOGRAFTS
*RNA
Subjects
Details
- Language :
- English
- ISSN :
- 00222836
- Volume :
- 329
- Issue :
- 4
- Database :
- Academic Search Index
- Journal :
- Journal of Molecular Biology
- Publication Type :
- Academic Journal
- Accession number :
- 9918594
- Full Text :
- https://doi.org/10.1016/S0022-2836(03)00403-0