Two-dimensional electrophoresis of Malassezia allergens for atopic dermatitis and isolation of Mal f 4 homologs with mitochondrial malate dehydrogenase.
The yeast Malassezia furfur is a natural inhabitant of the human skin microflora that induces an allergic reaction in atopic dermatitis. To identify allergens of M. furfur, we separated a crude preparation of M. furfur antigens as discrete spots by 2-D PAGE and detected IgE-binding proteins using sera of atopic dermatitis patients. We identified the known allergens, Mal f 2 and Mal f 3, and determined N-terminal amino acid sequences of six new IgE-binding proteins including Mal f 4. The cDNA and genomic DNA encoding Mal f 4 were cloned and sequenced. The gene was mitochondrial malate dehydrogenase and encoded Mal f 4 composed of 315 amino acids and a signal sequence of 27 amino acids. We purified Mal f 4, which had a molecular mass of 35 kDa from a membrane fraction of a lysate of cultured cells. Thirty of 36 M. furfur-allergic atopic dermatitis patients (83.3%) had elevated serum levels of IgE to purified Mal f 4, indicating that Mal f 4 is a major allergen. There was a significant correlation of the Phadebas RAST unit values of Mal f 4 and the crude antigen, but not between Mal f 4 and the known allergen Mal f 2.
Onishi
Y
Y
Biotechnology Research Laboratories, Takara Shuzo Co. Ltd, Shiga, Japan.
Kuroda
M
M
Yasueda
H
H
Saito
A
A
Sono-Koyama
E
E
Tunasawa
S
S
Hashida-Okado
T
T
Yagihara
T
T
Uchida
K
K
Yamaguchi
H
H
Akiyama
K
K
Kato
I
I
Takesako
K
K
D000485:Allergens / Q000235:genetics / Q000302:isolation & purification*; D000595:Amino Acid Sequence; D000946:Antigens, Fungal / Q000235:genetics / Q000302:isolation & purification; D052179:Antigens, Plant; D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D003876:Dermatitis, Atopic / Q000276:immunology*; D015180:Electrophoresis, Gel, Two-Dimensional; D005800:Genes, Fungal; D006801:Humans; D007073:Immunoglobulin E / Q000097:blood; D008290:Malassezia / Q000201:enzymology / Q000235:genetics / Q000276:immunology*; D008291:Malate Dehydrogenase / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data
D000485:Allergens; D000946:Antigens, Fungal; D052179:Antigens, Plant; D018076:DNA, Complementary; D004271:DNA, Fungal; C118831:Mal f 4 allergen; D007073:Immunoglobulin E; D008291:Malate Dehydrogenase