Vacuolar Serine Protease Is a Major Allergen of <i>Cladosporium cladosporioides</i>

Abstract

Background:Cladosporium is an important allergenic fungus worldwide. We report here a major allergen of C. cladosporioides.Methods: Major C. cladosporioides allergens were characterized by immunoblotting, N-terminal amino acid sequencing, protein purification and cDNA cloning. Results: Seventy-four sera (38%) from 197 bronchial asthmatic patients demonstrated IgE binding against C. cladosporioides extracts. Among these 74 sera, 41 (55%) and 38 (51%) showed IgE binding against a 36- and a 20-kDa protein of C. cladosporioides, respectively. Both IgE-reacting components reacted with FUM20, a monoclonal antibody against fungal serine proteases. N-terminal amino acid sequencing results suggest that they are vacuolar serine proteases, and the 20-kDa component is possibly a degraded product of the 36-kDa allergen. A corresponding 5′-truncated 1,425-bp cDNA fragment was isolated. The mature protein after N-terminal processing starts with an N-terminal serine that is the ninth residue encoded by the 5′-truncated cDNA. The protein sequence deduced shares 69–72% sequence identity with Penicillium vacuolar serine proteases and was designated as Cla c 9. The purified 36-kDa Cla c 9 allergen showed proteolytic activity with peptide Z-Ala-Ala-Leu-pNA as substrate. IgE cross-reactivity was detected between the purified Cla c 9 and serine protease allergens from Aspergillusfumigatus and Penicillium chrysogenum.Conclusion: We identified a vacuolar serine protease as a major allergen of C. cladosporioides (Cla c 9) and a major pan-allergen of prevalent airborne fungi. IgE cross-reactivity among these highly conserved serine protease pan-fungal allergens was also detectable.

The Transaldolase, a Novel Allergen of Fusarium proliferatum, Demonstrates IgE Cross-Reactivity with Its Human Analogue

“…The CYB medium contains yeast carbon base (Difco Laboratories, Detroit MI, USA; 11.7 g/L), glucose (Mallinckrodt Baker, Inc., Phillipsburg, NJ, USA; 10 g/L) and casein enzymatic hydrolysate (Sigma Chemical Co., St. Louis, MO, USA; 10 g/L). Crude extracts of F. proliferatum were prepared essentially as described [19], [20]. The protein content of crude fungal extracts was determined with a dye-binding assay according to the manufacturer’s instructions (Bio-Rad, Richmond, CA, USA).…”

“…Proteins in the crude fungal extractions or purified recombinant proteins were separated by SDS-PAGE [19], [20] then transferred electrophoretically onto polyvinylidene difluoride (PVDF) membranes (0.45 µm, Millipore, Bedford, MS, USA). Protein components reacting against human IgE antibodies were determined as described [19], [20].…”

Molecular and immunological characterization of subtilisin like serine protease, a major allergen of Curvularia lunata

“…Proteolytic activity increases bronchial epithelial permeability by degrading proteins of desmosomes, tight and adherens junctions and thereby facilitating antigen access to the immune system (Robinson et al, 2001). Fungal serine proteases such as Asp fl 1, Cla c 9, Cla h 9, Pen ch 13, Pen ch 18 and Pen o 18 were cloned and expressed as recombinant proteins (Yu et al, 1999;Shen et al, 2001Shen et al, , 2003Chou et al, 2002Chou et al, , 2008Pöll et al, 2009). These homogeneous preparations can be modified by reducing allergenic activity to foster advantageous immunologic properties like abrogating IgE binding epitopes or tagging novel sequences for obtaining better immunotherapeutic response.…”

Free-Living Amoeba Acanthamoeba Triggers Allergic Inflammation of Airways

“…Ovalbumin (OVA) is one of commonly used the experimental allergens, but is unable to elicit allergic airway inflammation if directly administered by inhalation without any allergens. By contrast, pollens and fungal-derived allergens can easily elicit allergic responses when inhaled through the airway tract [17][18][19]. Therefore, if repeated administration of Acanthamoeba into the airway tract, allergic response can be occurred in the airway.…”