Novel mite allergen

Abstract

A safe and efficient recombinant mite allergen is provided as a therapeutic agent or a diagnostic agent for mite allergic diseases, which contains no anaphylaxis-inducing impurities. The following recombinant protein (a) or (b) is provided: (a) a protein comprising the amino acid sequence represented by SEQ ID NO: 2 or 35; or (b) a protein comprising an amino acid sequence derived from the amino acid sequence represented by SEQ ID NO: 2 or 35 by deletion, substitution, or addition of one or several amino acids and having mite allergen activity.

What is claimed is: 
     
         1 . A polypeptide fragment of a mite allergen derived from  Dermatophagoides farinae  comprising any one of SEQ ID NOs: 2, 4, 10, 12, or 19. 
     
     
         2 . A therapeutic agent for mite allergic diseases, comprising a composition having the polypeptide fragment according to  claim 1  as an active ingredient. 
     
     
         3 . A diagnostic agent for mite allergic diseases, wherein the diagnostic agent comprises the polypeptide fragment according to  claim 1 . 
     
     
         4 . A polypeptide fragment of a mite allergen derived from  Dermatophagoides farinae  comprising SEQ ID NOs: 4, 10, 12, and 19. 
     
     
         5 . A therapeutic agent for mite allergic diseases, comprising a composition having the polypeptide fragment according to  claim 4  as an active ingredient. 
     
     
         6 . A diagnostic agent for mite allergic diseases, wherein the diagnostic agent comprises the polypeptide fragment according to  claim 4 . 
     
     
         7 . A mite allergen, which is obtained by:
 (i) extracting mite poly(A) mRNA from the mite bodies of  Dermatophagoides farinae;      (ii) producing cDNAs by reverse transcription using the mite poly(A) mRNA of (i);   (iii) amplifying a gene by PCR using primer N-1 (5′-GAYGAYGTNTTRAARCARACNGARGAR-3′ (SEQ ID NO: 20): Y=C or T, N=A or C or G or T, and R=A or G) as a sense primer and a primer having the nucleotide sequence of SEQ ID NO: 25 as a reverse primer, and using the mite cDNA obtained in (ii) as a template;   (iv) introducing the DNA fragment obtained in (iii) into an expression vector and transforming host  Escherichia coli  with the vector;   (v) producing a protein which has the molecular weight of 150 kDa to 250 kDa and an N-terminal sequence comprising SEQ ID NO: 19 by culturing the host  Escherichia coli  obtained in (iv);   (vi) immunizing a mouse with the protein obtained in (v);   (vii) obtaining a polyclonal antibody against the mite allergen from the immunized mouse of (vi);   (viii) allowing the polyclonal antibody obtained in (vii) to come into contact with antigens extracted from  Dermatophagoides farinae ; and   (ix) collecting an antigen which is bound to the polyclonal antibody obtained in (vii).   
     
     
         8 . The mite allergen according to  claim 7 , in which the amino acid sequence PEPTTKT (amino acids 131-137 of SEQ ID NO: 35) is repeated at least 11 times.