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Recombinant lipid transfer protein Tri a 14: a novel heat and proteolytic resistant tool for the diagnosis of baker's asthma.
Clin Exp Allergy 2009; 39(8):1267-76CE

Abstract

BACKGROUND

Baker's asthma is an important occupational allergic disease. Wheat lipid transfer protein (LTP) Tri a 14 is a major allergen associated with wheat allergy. No panel of wheat recombinant allergens for component-resolved diagnosis of baker's asthma is currently available.

OBJECTIVE

To evaluate the potential role of recombinant Tri a 14 as a novel tool for the diagnosis of baker's asthma, and to test the heat and proteolytic resistance of the wheat LTP allergen.

METHODS

A cDNA encoding Tri a 14 was isolated and sequenced, the recombinant allergen produced in Pichia pastoris and purified by chromatographic methods. Physicochemical and immunological comparison of the natural and recombinant forms of Tri a 14 was carried out by N-terminal amino acid sequencing, matrix-assisted laser desorption/ionization mass spectrometry, circular dichroism (CD) analysis, IgE immunodetection, and specific IgE determination and ELISA-inhibition assays using a pool or individual sera from 26 patients with baker's asthma. Thermal denaturation and simulated gastrointestinal digestion of both Tri a 14 forms were checked by spectroscopic and electrophoretic methods, respectively, and biological activity by basophil activation test (BAT).

RESULTS

Natural and recombinant Tri a 14 were similarly folded, as indicated by their nearly identical CD spectra and heat denaturation profiles. A high interclass correlation coefficient (0.882) was found between specific IgE levels to both Tri a 14 proteins in individual sera from baker's asthma patients, but a slightly lower IgE-binding potency of rTri a 14 was detected by ELISA-inhibition assays. Natural and recombinant Tri a 14 elicited positive BAT in two and one out of three patients, respectively. Heat denaturation profiles and simulated gastrointestinal digestion assays indicated that Tri a 14 displayed a high heat and digestive proteolytic resistance, comparable to those of peach Pru p 3, the model food allergen of the LTP family.

CONCLUSIONS

Recombinant Tri a 14 is a potential tool for baker's asthma diagnosis, based on its physicochemical and immunological similarity with its natural counterpart. Wheat Tri a 14 shows a high thermal stability and resistance to gastrointestinal digestion.

Authors+Show Affiliations

Unidad de Bioquimica, Departamento de Biotecnología, E.T.S. Ingenieros Agronomos, Madrid, Spain.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

19486028

Citation

Palacin, A, et al. "Recombinant Lipid Transfer Protein Tri a 14: a Novel Heat and Proteolytic Resistant Tool for the Diagnosis of Baker's Asthma." Clinical and Experimental Allergy : Journal of the British Society for Allergy and Clinical Immunology, vol. 39, no. 8, 2009, pp. 1267-76.
Palacin A, Varela J, Quirce S, et al. Recombinant lipid transfer protein Tri a 14: a novel heat and proteolytic resistant tool for the diagnosis of baker's asthma. Clin Exp Allergy. 2009;39(8):1267-76.
Palacin, A., Varela, J., Quirce, S., del Pozo, V., Tordesillas, L., Barranco, P., ... Salcedo, G. (2009). Recombinant lipid transfer protein Tri a 14: a novel heat and proteolytic resistant tool for the diagnosis of baker's asthma. Clinical and Experimental Allergy : Journal of the British Society for Allergy and Clinical Immunology, 39(8), pp. 1267-76. doi:10.1111/j.1365-2222.2009.03280.x.
Palacin A, et al. Recombinant Lipid Transfer Protein Tri a 14: a Novel Heat and Proteolytic Resistant Tool for the Diagnosis of Baker's Asthma. Clin Exp Allergy. 2009;39(8):1267-76. PubMed PMID: 19486028.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Recombinant lipid transfer protein Tri a 14: a novel heat and proteolytic resistant tool for the diagnosis of baker's asthma. AU - Palacin,A, AU - Varela,J, AU - Quirce,S, AU - del Pozo,V, AU - Tordesillas,L, AU - Barranco,P, AU - Fernandez-Nieto,M, AU - Sastre,J, AU - Diaz-Perales,A, AU - Salcedo,G, Y1 - 2009/05/26/ PY - 2009/6/3/entrez PY - 2009/6/3/pubmed PY - 2009/12/18/medline SP - 1267 EP - 76 JF - Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology JO - Clin. Exp. Allergy VL - 39 IS - 8 N2 - BACKGROUND: Baker's asthma is an important occupational allergic disease. Wheat lipid transfer protein (LTP) Tri a 14 is a major allergen associated with wheat allergy. No panel of wheat recombinant allergens for component-resolved diagnosis of baker's asthma is currently available. OBJECTIVE: To evaluate the potential role of recombinant Tri a 14 as a novel tool for the diagnosis of baker's asthma, and to test the heat and proteolytic resistance of the wheat LTP allergen. METHODS: A cDNA encoding Tri a 14 was isolated and sequenced, the recombinant allergen produced in Pichia pastoris and purified by chromatographic methods. Physicochemical and immunological comparison of the natural and recombinant forms of Tri a 14 was carried out by N-terminal amino acid sequencing, matrix-assisted laser desorption/ionization mass spectrometry, circular dichroism (CD) analysis, IgE immunodetection, and specific IgE determination and ELISA-inhibition assays using a pool or individual sera from 26 patients with baker's asthma. Thermal denaturation and simulated gastrointestinal digestion of both Tri a 14 forms were checked by spectroscopic and electrophoretic methods, respectively, and biological activity by basophil activation test (BAT). RESULTS: Natural and recombinant Tri a 14 were similarly folded, as indicated by their nearly identical CD spectra and heat denaturation profiles. A high interclass correlation coefficient (0.882) was found between specific IgE levels to both Tri a 14 proteins in individual sera from baker's asthma patients, but a slightly lower IgE-binding potency of rTri a 14 was detected by ELISA-inhibition assays. Natural and recombinant Tri a 14 elicited positive BAT in two and one out of three patients, respectively. Heat denaturation profiles and simulated gastrointestinal digestion assays indicated that Tri a 14 displayed a high heat and digestive proteolytic resistance, comparable to those of peach Pru p 3, the model food allergen of the LTP family. CONCLUSIONS: Recombinant Tri a 14 is a potential tool for baker's asthma diagnosis, based on its physicochemical and immunological similarity with its natural counterpart. Wheat Tri a 14 shows a high thermal stability and resistance to gastrointestinal digestion. SN - 1365-2222 UR - https://www.unboundmedicine.com/medline/citation/19486028/Recombinant_lipid_transfer_protein_Tri_a_14:_a_novel_heat_and_proteolytic_resistant_tool_for_the_diagnosis_of_baker's_asthma_ L2 - https://doi.org/10.1111/j.1365-2222.2009.03280.x DB - PRIME DP - Unbound Medicine ER -