Characterization of a novel Candida albicans 29 kDa IgE-binding protein - purification, cDNA isolation and heterologous expression of Cand a 3 (original) (raw)
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Extraction and Purification of the Potential Allergen Proteins from Candida Albicans
DergiPark (Istanbul University), 2020
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Mycoses, 2009
Enzyme-immunoassay-Candida albicans-Mannan-antigen-Ig-Isotypes Schliisselworter: Enzym-Immunoassay-Candida albicans-Mannan-Antigen-Ig-Isotypen Summary: The serum levels of anti-Candida albicans IgA, IgG and IgM class antibodies were quantified by heterogenous sandwich enzyme-immunoassay. The presence of polysaccharide antigen reactive with rabbit antiserum to Candida albicans was also demonstrated by this assay. The thermal dissociation of antigen-antibody complex was proved to be efficient in quantitative determination of fungal antigens. Using the methods of correlation analysis, the close relationship between Ig-isotypes was studied. The statistically significant correlation between IgG and IgA class (rX,,=0.475), and IgA and IgM class (r,,,=0.465) was revealed. The correlation between IgG and IgM class (r,,=0375) for a=O.O1 was decreased signifcantly. Zusammenfassung: Die Serumspiegel der Anti-Candida albicans Aatikorper der Klassen IgA, IgG und IgM wurden durch den heterogenen Sandwich-Enzymimmunoassay quantifiiiert. Die Anwesenheit von Polysaccharidantigen, das mit Kaninchenantiserum gegen Candida albicans reagiert, wurde ebenfalls durch diesen Assay dargestellt. Die Hike-Dissoziation des Antigen-Antikiirperkomplexes envies sich bei der quantitativen Bestimmung des Phantigens als wirksam. Unter Verwendung der Methoden der Korrelationsanalyse wurde die enge Beziehung zwischen Ig-Isotypen studiert. Es zeigte sich eine statistisch significante Korrelation zwischen IgG und IgA Klasse (r,,=0.475) und IgA und IgM Klasse (r,,=0.465). Die Korrelationzwischen IgG und IgM Klasse war signifikant abnehmend (rX,,=0375) auf a=O.Ol.
IgE-binding epitopes of enolases, a class of highly conserved fungal allergens
Journal of Allergy and Clinical Immunology, 2000
Background: Cladosporium herbarum and Alternaria alternata are two of the most prominent fungal species inducing type I allergy. Previously, we have demonstrated that enolase (Cla h 6) is the second most important allergen of C herbarum in terms of frequency of sensitization. Objective: IgE-reactive B-cell epitopes of C herbarum enolase were analyzed, and cross-reactivity between fungal enolases was investigated. Methods: Cla h 6 glutathione-S-transferase fusion peptides were constructed by means of PCR cloning. A alternata enolase (Alt a 5) was isolated by screening a complementary (c)DNA expression library with a C herbarum enolase DNA probe. Results: Mapping of Cla h 6 IgE-binding epitopes identified a peptide with a length of 69 amino acids (peptide 9), which bound IgE from 8 of 8 patients. Analysis of the conformation of peptide 9 revealed that it does not form a compact structure but rather spans the whole length of the protein, with side chains exposed to solvent at 3 locations. Peptide 9 in the context of Escherichia coli glutathione-S-transferase not only binds IgE but also competitively inhibits IgE binding to Alt a 5. This result indicates that the epitope or epitopes on peptide 9 constitute a major cross-reacting epitope or epitopes on the enolases from C herbarum and A alternata in the case of the one patient tested. Conclusions: We demonstrated that the glycolytic enzyme enolase is an allergen not only in C herbarum but also in A alternata. Additionally, enolase was shown to exhibit high crossreactivity to other fungal enolases. On the basis of the results presented here, we propose the use of recombinant Cla h 6 or maybe even peptide 9 of Cla h 6 for diagnosis and possibly therapy of mold allergy. (J Allergy Clin Immunol 2000;106: 887-95.)
Identification of antigens reacting with anti-Candida albicans germ tube antibodies
European Journal of Epidemiology, 1992
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Data in brief, 2016
The characterization of pathogen-specific antigenic proteins at the protein species level is crucial in the development and molecular optimization of novel immunodiagnostics, vaccines or immunotherapeutics for infectious diseases. The major requirements to achieve this molecular level are to obtain 100% sequence coverage and identify all post-translational modifications of each antigenic protein species. In this article, we show nearly complete sequence information for five discrete antigenic species of Candida albicans Tdh3 (glyceraldehyde-3-phosphate dehydrogenase), which have been reported to be differentially recognized both among candidemia patients and between candidemia and control patients. A comprehensive description of the top-down immunoproteomic strategy used for seroprofiling at the C. albicans protein species level in candidemia as well as for the chemical characterization of this immunogenic protein (based on high-resolution 2-DE, Western blotting, peptide mass finger...
Novel Cytosolic Allergens of Aspergillus fumigatus Identified from Germinating Conidia
Journal of Proteome Research, 2010
Aspergillus fumigatus is the common cause of allergic broncho-pulmonary aspergillosis (ABPA) and most of the allergens have been described from its secreted fraction. In the present investigation, germinating conidial cytosolic proteins of A. fumigatus were extracted from a 16 h culture. The proteome from this fraction was developed, and immuno-blots were generated using pooled ABPA patients' sera. Well separated Immunoglobulin-E (IgE) and Immunoglobulin-G (IgG) reactive spots were picked from corresponding 2DE gels and subjected to mass spectrometric analysis. As a result, 66 immuno-reactive proteins were identified from two geographically different strains (190/96 and DAYA) of A. fumigatus. Only 3 out of 66 proteins reacted with IgG, and the remaining 63 proteins were found to be IgE reactive. These 63 IgE-reactive cytosolic proteins from germinating conidia included 2 already known (Asp f12 and Asp f22) and 4 predicted allergens (Hsp88, Hsp70, malate dehydrogenase, and alcohol dehydrogenase) based on their homology with other known fungal allergens. In view of this, the panel of presently identified IgE-reactive novel proteins holds the potential of providing a basis for the wider diagnostic application in assay for allergic aspergillosis. We could demonstrate that recombinantly expressed proteins from this panel showed consistent reactivity with IgE of individual sera of ABPA patients. The recombinantly expressed proteins may also be useful in desensitization therapy of allergic disorders including ABPA.
Revista Iberoamericana De Micologia
Background: Candida albicans is an opportunistic dimorphic fungus commonly present in the human oral cavity that causes infections in immunocompromised patients. The antigen variability, influenced by growth conditions, is a pathogenicity factor. Aims: To determine the effect of nutritional and heat stress on the antigen expression of C. albicans, and to identify major antigens recognized by human salivary secretory immunoglobulin A (sIgA). Methods: Under various different nutritional conditions, heat shock was induced in C. albicans cells in stationary and exponential growth phases. The expression of protein determinants of C. albicans was assessed by Western blot analysis against human saliva. The antigens were purified and characterized by two-dimensional electrophoresis and identified by protein microsequencing. Results: Five antigens recognized by salivary IgA were characterized as mannoproteins due to their reactivity with concanavalin A. They did not show reactivity with anti-heat shock protein monoclonal antibodies. Two of them (42 and 36 kDa) were found to be regulated by heat shock and by nutritional stress and they were identified as phosphoglycerate kinase and fructose bisphosphate aldolase, respectively. Conclusions: These glycolytic enzymes are major antigens of C. albicans, and their differential expression and recognition by the mucosal immune response system could be involved in protection against oral infection.