title: Molecular characterization of the group 4 house dust mite allergen from Dermatophagoides pteronyssinus and its amylase homologue from Euroglyphus maynei.
abstract: BACKGROUND
  Of the ten recognised groups of Dermatophagoides pteronyssinus allergens, the group 4 is the only group that has not been characterised at the molecular level.
  
  
  METHODS
  Primers were designed to PCR amplify Der p 4 (D. pteronyssinus) and Eur m 4 (Euroglyphus maynei) cDNA. These fragments were used to screen the corresponding cDNA libraries and the cDNA clones obtained were subsequently sequenced. The coding regions of Der p 4 and Eur m 4 were cloned into the pET expression vector and recombinant histidine-tagged proteins expressed in Escherichia coli.
  
  
  RESULTS
  cDNA clones which included the mature protein coding sequence for Der p 4 and Eur m 4 were sequenced. The Der p 4 and Eur m 4 genes were found to code for 496 amino acid mature proteins with residues important for the function of alpha-amylase highly conserved. Der p 4 and Eur m 4 were calculated to be 90% identical and a BLAST search of the GenBank database found these sequences to be approximately 50% identical to insect and mammalian alpha-amylases. The calculated molecular weights of Der p 4 and Eur m 4 were approximately 57,000, although recombinant Der p 4 and Eur m 4 migrate on SDS-PAGE at about 60,000. Der p 4 recombinant protein was found to bind specific IgE in 3 of the 10 house dust mite allergic patients tested.
  
  
  CONCLUSIONS
  This paper describes the first cDNA sequence of Der p 4 and Eur m 4 confirming that this allergen is house dust mite alpha-amylase.
authors.affiliation: TVW Telethon Institute for Child Health Research, West Perth, Australia.
authors.forename: K L
authors.initials: KL
authors.lastname: Mills
authors.forename: B J
authors.initials: BJ
authors.lastname: Hart
authors.forename: N R
authors.initials: NR
authors.lastname: Lynch
authors.forename: W R
authors.initials: WR
authors.lastname: Thomas
authors.forename: W
authors.initials: W
authors.lastname: Smith
mesh_terms: D000485:Allergens / Q000276:immunology; D000595:Amino Acid Sequence; D000681:Amylases / Q000235:genetics; D000818:Animals; D039741:Antigens, Dermatophagoides; D060829:Arthropod Proteins; D001483:Base Sequence; D004391:Dust / Q000032:analysis; D015723:Gene Library; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics*; D019476:Insect Proteins; D008925:Mites / Q000276:immunology; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D017386:Sequence Homology, Amino Acid
chemical_list: D000485:Allergens; D039741:Antigens, Dermatophagoides; D060829:Arthropod Proteins; D004391:Dust; C079269:Eur m I protein, Euroglyphus maynei; D006023:Glycoproteins; D019476:Insect Proteins; D011994:Recombinant Proteins; D000681:Amylases