Characterization of Der f 22 - a paralogue of the major allergen Der f 2

Reginald, Kavita; Tan, Chew-Lim; Chen, Simin; Yuen, Liling; Goh, Sock Yong; Chew, Fook Tim

doi:10.1038/s41598-018-30224-z

Cited by 6 publications

(4 citation statements)

References 29 publications

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“…Using well established lipid binding assays and mass spectrometry, we show direct evidence that Der p 2 is a cholesterol binding protein. In addition, we also show evidence that homologues of Der p 2, specifically Der f 2 and Der f 22, a paralogue of Der f 2 12 , binds to cholesterol.…”

Section: Introductionsupporting

confidence: 57%

“…While group 2 allergens may bind to a range of lipid molecules, our data clearly shows that it shows preferential binding to cholesterol. In addition, two other allergens with structural similarities to Der p 2, namely Der f 2, a well characterized group 2 allergen from D. farinae and Der f 22, a paralogue of group 2 allergens 12 also bind to cholesterol in a dose-responsive manner, similar to Der p 2.…”

Section: Discussionmentioning

confidence: 99%

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The major allergen Der p 2 is a cholesterol binding protein

Reginald

Chew

2019

Sci Rep

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Der p 2 is a major dust mite allergen and >80% of mite allergic individuals have specific IgE to this allergen. Although it is well characterized in terms of allergenicity, there is still some ambiguity in terms of its biological function. Three-dimensional structural analysis of Der p 2 and its close homologues indicate the presence of a hydrophobic cavity which can potentially bind to lipid molecules. In this study, we aimed to identify the potential ligand of Der p 2. Using a liposome pulldown assay, we show that recombinant Der p 2 binds to liposomes prepared with exogenous cholesterol in a dose dependent fashion. Next, an ELISA based assay using immobilized lipids was used to study binding specificities of other lipid molecules. Cholesterol was the preferred ligand of Der p 2 among 11 different lipids tested. Two homologues of Der p 2, Der f 2 and Der f 22 also bound to cholesterol. Further, using liquid chromatography-mass spectrometry (LC-MS), we confirmed that cholesterol is the natural ligand of Der p 2. Three amino acid residues of Der p 2, V104, V106 and V110 are possible cholesterol binding sites, as alanine mutations of these residues showed a significant decrease in binding (p < 0.05) compared to wild-type Der p 2. These results provide the first direct experimental evidence that Der p 2 binds to cholesterol.

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Section: Introductionsupporting

confidence: 57%

Section: Discussionmentioning

confidence: 99%

The major allergen Der p 2 is a cholesterol binding protein

Reginald

Chew

2019

Sci Rep

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“…Immuno-dot blot assays were performed as previously reported 64 . Serially diluted IgE standards (National Institute for Biological Standards, UK) and bovine serum albumin (BSA) were used as a positive and negative controls respectively.…”

Section: Methodsmentioning

confidence: 99%

Blo t 2: Group 2 allergen from the dust mite Blomia tropicalis

Reginald

Pang

Chew

2019

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Blomia tropicalis has been recognized as a cause of allergic diseases in the tropical and subtropical regions. Here we report the immuno-characterization of its group 2 allergen, Blo t 2. Allergen Blo t 2 was amplified from the cDNA of B. tropicalis using degenerate primers, expressed in Escherichia coli as a recombinant protein and purified to homogeneity. The mature protein of Blo t 2 was 126 amino acids long with 52% sequence identity to Der p 2 and apparent molecular mass of 15 kDa. Circular dichroism spectroscopy showed that Blo t 2 is mainly a beta-sheeted protein. We confirmed the presence of three disulfide bonds in recombinant (r) Blo t 2 protein using electrospray mass spectrometry. Thirty-four percent of dust-mite allergic individuals from the Singapore showed specific IgE binding to rBlo t 2 as tested using immuno dot-blots. IgE-cross reactivity assays showed that Blo t 2 had between 20–50% of unique IgE-epitopes compared to Der p 2. IgE binding of native and recombinant forms of Blo t 2 were highly concordant (r 2 = 0.77, p < 0.0001) to rBlo t 2. Dose-dependent in vitro histamine was observed when rBlo t 2 was incubated with whole blood of Blo t 2 sensitized individuals, demonstrating that it is a functional allergen. Nine naturally occurring isoforms of Blo t 2 were identified in this study, each having between 1–3 amino acid variations compared to the reference clone. Blo t 2 is a clinically relevant allergen of B. tropicalis as it has unique IgE epitopes compared to major group 2 allergens from Dermatophagoides spp .

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“…1B). Considering that group 22 and 35 allergens were both characterized as members of the NPC2 family 24-26 , we will perform a comparative analysis in astigmatic mites to further explore this important gene family.…”

Section: Resultsmentioning

confidence: 99%

Multi-Omic Analysis ofTyrophagus putrescentiaeReveals Insights into the Allergen Complexity of Storage Mites

Wan

Xiong

Xiao

et al. 2022

Preprint

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BackgroundThe storage miteTyrophagus putrescentiaeis one of the major mites causing allergies in Chinese and Korean populations, but its allergen profile in incomplete when compared with that of house dust mites. Multiple genome-based methods have been introduced into the allergen study of mites and have enabled a better understanding of these medically important organisms.ObjectiveWe sought to reveal a comprehensive allergen profile ofTyrophagus putrescentiaeand advance the allergen study of storage mites.MethodsBased on a high-quality assembled and annotated genome, anin silicoanalysis was performed by searching reference sequences to identify allergens. Immunoassay ELISA assessed the allergenicities of recombinant proteins. MALDI-TOF mass spectrometry identified the IgE-binding proteins. Comparative genomics analysis was employed for the important allergen gene families.ResultsA complete allergen profile ofTyrophagus putrescentiaewas revealed, including thirty-seven allergen groups (up to Tyr p 42). Among them, five novel allergens were verified using the sera of allergy patients. Massive allergen homologs were identified as the result of gene duplications in genome evolution. Proteomic identification again revealed a wide range of allergen homologs. In the NPC2 family and GSTs, comparative analysis shed light on the expansion and diversification of the allergen groups.ConclusionUsing multi-omic approaches, the comprehensive allergen profile including massive homologs was disclosed inTyrophagus putrescentiae, which revealed the allergen complexity of the storage mite and could ultimately facilitate the component-resolved diagnosis.

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Characterization of Der f 22 - a paralogue of the major allergen Der f 2

Cited by 6 publications

References 29 publications

The major allergen Der p 2 is a cholesterol binding protein

The major allergen Der p 2 is a cholesterol binding protein

Blo t 2: Group 2 allergen from the dust mite Blomia tropicalis

Multi-Omic Analysis ofTyrophagus putrescentiaeReveals Insights into the Allergen Complexity of Storage Mites

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