NADP-dependent mannitol dehydrogenase, a major allergen of Cladosporium herbarum

J Biol Chem. 2006 Jun 16;281(24):16354-60. doi: 10.1074/jbc.M513638200. Epub 2006 Apr 11.

Abstract

Cladosporium herbarum is an important allergenic fungal species that has been reported to cause allergic diseases in nearly all climatic zones. 5-30% of the allergic population displays IgE antibodies against molds. Sensitization to Cladosporium has often been associated with severe asthma and less frequently with chronic urticaria and atopic eczema. However, no dominant major allergen of this species has been found so far. We present cloning, production, and characterization of NADP-dependent mannitol dehydrogenase of C. herbarum (Cla h 8) and show that this protein is a major allergen that is recognized by IgE antibodies of approximately 57% of all Cladosporium allergic patients. This is the highest percentage of patients reacting with any Cladosporium allergen characterized so far. Cla h 8 was purified to homogeneity by standard chromatographic methods, and both N-terminal and internal amino acid sequences of protein fragments were determined. Enzymatic analysis of the purified natural protein revealed that this allergen represents a NADP-dependent mannitol dehydrogenase that interconverts mannitol and d-fructose. It is a soluble, non-glycosylated cytoplasmic protein. Two-dimensional protein analysis indicated that mannitol dehydrogenase is present as a single isoform. The cDNA encoding Cla h 8 was cloned from a lambda-ZAP library constructed from hyphae and spores. The recombinant non-fusion protein was expressed in Escherichia coli and purified to homogeneity. Its immunological and biochemical identity with the natural protein was shown by enzyme activity tests, CD spectroscopy, IgE immunoblots with sera of patients, and by skin prick testing of Cladosporium allergic patients. This protein therefore is a new major allergen of C. herbarum.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alcohol Oxidoreductases / metabolism*
  • Allergens / chemistry
  • Amino Acid Sequence
  • Base Sequence
  • Circular Dichroism
  • Cladosporium / chemistry
  • Cladosporium / immunology*
  • Cytoplasm / metabolism
  • Electrophoresis, Gel, Two-Dimensional
  • Humans
  • Immunoglobulin E / chemistry
  • Mannitol Dehydrogenases
  • Molecular Sequence Data
  • Oligonucleotides / chemistry
  • Protein Structure, Tertiary
  • Skin Tests

Substances

  • Allergens
  • Oligonucleotides
  • Immunoglobulin E
  • Alcohol Oxidoreductases
  • Mannitol Dehydrogenases
  • mannitol 2-dehydrogenase (NADP)

Associated data

  • GENBANK/AY191816