Cloning, expression and characterization of two new IgE‐binding proteins from the yeast Malassezia sympodialis with sequence similarities to heat shock proteins and manganese superoxide dismutase

Abstract

Malassezia sympodialis is an opportunistic yeast that colonizes human skin and may induce IgE and T cell reactivity in patients with atopic eczema/dermatitis syndrome (AEDS). Previously, we have cloned and expressed six recombinant allergens (rMala s 1 and rMala s 5 to rMala s 9) from this yeast. By combining high throughput screening and phage surface display techniques, 27 complete and partial IgE-binding clones of M. sympodialis have been identified. Here we enlarged the panel of recombinant M. sympodialis allergens by RACE-PCR, cloning and nucleotide sequencing to obtain the coding sequences of two new IgE-binding clones. The coding sequences of one of the clones showed similarity to the heat shock protein (HSP) family and the other to manganese superoxide dismutase (MnSOD), and both had a high degree of homology to human counterparts. The coding sequences were expressed in Escherichia coli as six-histidine tagged recombinant proteins and generated products with molecular masses of 86.1 kDa for HSP and 22.4 kDa for MnSOD. Their IgE-binding frequencies were shown to be 69% and 75%, respectively, to 28 sera from AEDS patients with serum IgE to M. sympodialis extract, indicating that HSP and MnSOD are major M. sympodialis allergens. In inhibition immunoblotting, M. sympodialis extract could inhibit the binding of serum IgE from AEDS patients to rHSP and rMnSOD in a concentrationdependent manner. The high frequency of sera from AEDS patients, showing IgE binding to both HSP and MnSOD, indicates that these allergens, designated Mala s 10 and Mala s 11, could play a role in AEDS.

The <i>Malassezia Sympodialis </i>Allergen Mala s 11 Induces Human Dendritic Cell Maturation, in Contrast to Its Human Homologue Manganese Superoxide Dismutase

“…Its antioxidant activity protects the skin against the toxicity of oxygen-related radicals and the enzyme is up-regulated by UV exposure, mechanical trauma [20], and in the lesional skin of patients with AE [11]. The 2 proteins chosen are of particular interest, since sensitization to Mala s 11 has been observed in AE patients [7] and hMnSOD has been suggested to be self-reactive in these patients [11]. The results of the present study suggest that Mala s 11 can affect the immune system through DCs with the up-regulation of the maturation marker CD83, the co-stimulatory molecules CD40, CD80, CD86 and HLA-DR necessary for antigen stimulation to T cells.…”

“…Interestingly, Mala s 11 has a high degree of sequence homology to manganese superoxide dismutase (MnSOD) from Homo sapiens (50%) and Aspergillus fumigatus (56%) [7]. It has been observed that MnSOD derived from H. sapiens is able to bind IgE in sera from asthmatic patients allergic to A. fumigatus [8].…”

“…The Mala s 11 allergen (22.4 kDa) is a major allergen with an IgE-binding frequency of 75% in sera from AE patients sensitized to M. sympodialis [7]. Interestingly, Mala s 11 has a high degree of sequence homology to manganese superoxide dismutase (MnSOD) from Homo sapiens (50%) and Aspergillus fumigatus (56%) [7].…”

Characterization of Cannabis Sativa Allergens

“…44 BiP also shares a high degree of sequence homology to heat shock protein 70 (Hsp70), which has been reported as an allergen in many fungi. 45,46 Oxygen evolving enhancer protein was a prominent allergen in our analysis; however, there are no studies in the literature that currently report this protein to be an allergen.…”

Cutaneous yeast microflora in patients with atopic dermatitis

“…These data can explain why fungi-allergic patients show reactivity to several fungal species [22]. Furthermore, there is evidence that the IgE against M. sympodialis protein manganese superoxide dismutase can cross-react with a corresponding human protein, leading to autoreactivity in patients with AD [23]. The majority of studies involving yeasts and AD patients focus mainly on the members of the genus Malassezia [24][25][26][27].…”