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Published on: February 24, 2021
Isolation, expression and characterization of a minor allergen from Penicillium crustosum
M Serdal Sevinc1, Veena Kumar, Makonnen Abebe
1Healthy Environments and Consumer Safety Branch, Health Canada, Ottawa, ON, Canada.
Abstract:
A ribosomal P1 protein, Pen b 26 from Penicillium brevicompactum, was previously identified as a major allergen. A homolog protein was isolated and characterized from Penicillium crustosum which is not known to be allergenic mold. A cDNA library of P. crustosum was constructed and screened using a probe based on the DNA sequence of Pen b 26. A positive clone was isolated, expressed in Escherichia coli, purified and characterized by comparing its immunological and physical properties to Pen b 26. It was designated as Pen cr 26 and had a 321 nt ORF corresponding to 107 amino acids with a MW of 11 kDa. Pen cr 26 had strong sequence homology to Pen b 26 (92% for nucleotides and 86% for amino acids) and its physical and predicted structural properties were similar to the latter. The level of expression of Pen cr 26 was much lower than that of Pen b 26 in the same expression vector. Both proteins were recognized equally well by the IgG class specific antibodies, but Pen cr 26 was poorly recognized by Penicillium-sensitive atopic sera (IgE), suggesting striking antigenic difference in IgE epitopes, i.e., 87% were positive for Pen b 26 while only 23% were positive for Pen cr 26. The allergenicity of Pen cr 26 seems to be minor in nature and it could be a hypoallergenic variant of Pen b 26.
Insights
A newly identified protein, Pen cr 26 from Penicillium crustosum, shows high similarity to the major allergen Pen b 26 but has significantly reduced allergenicity. This suggests Pen cr 26 may be a hypoallergenic variant.
Area of Science:
- Allergen research
- Molecular biology
- Mycology
Background:
- Penicillium species are common molds, with some, like Penicillium brevicompactum, producing major allergens such as Pen b 26.
- The allergenic potential of Penicillium crustosum, a related mold, is largely unknown.
- Understanding allergen variations can aid in managing mold allergies.
Purpose of the Study:
- To isolate and characterize a protein homologous to Pen b 26 from Penicillium crustosum.
- To compare the immunological and physical properties of the new protein with Pen b 26.
- To evaluate the allergenicity of the Penicillium crustosum homolog, designated Pen cr 26.
Main Methods:
- Construction and screening of a cDNA library from P. crustosum using a Pen b 26 DNA probe.
- Expression of the isolated clone in Escherichia coli, followed by purification.
- Characterization through immunological assays (IgG and IgE binding) and physical property comparison with Pen b 26.
Main Results:
- A homologous protein, Pen cr 26, was identified with high sequence similarity (92% nucleotide, 86% amino acid) to Pen b 26.
- Pen cr 26 exhibited similar physical and predicted structural properties to Pen b 26 but lower expression levels.
- While recognized by IgG antibodies, Pen cr 26 showed significantly reduced IgE binding (23% vs. 87% for Pen b 26) in allergic patient sera.
Conclusions:
- Pen cr 26 possesses distinct IgE epitopes compared to Pen b 26, leading to markedly lower allergenicity.
- The findings suggest Pen cr 26 is a minor allergen and potentially a hypoallergenic variant of Pen b 26.
- This research contributes to understanding allergen diversity within Penicillium species.
