Recombinant American cockroach component, Per a 1, reactive to IgE of allergic Thai patients.
Twelve similar recombinant Per a 1 clones were produced from an American cockroach (CR) cDNA library. The nucleotide sequence of a representative cline, i.e. clone A6, contained 579 base pairs (bp) and a 372 bp open reading frame (2-373) encoding 124 amino acids. A stop codon was found at position 374-376 followed by a 3' end untranslated region with an AATAAA polyadenylation signal and a poly (A) tail. The estimated molecular mass of the 24 amino acid residue protein was 13.8 kDa, with a predicted isoelectric point value of 4.74. Cysteine or N-linked glycosylation was not found. The deduced amino acid sequence of the A6 revealed 84.68-95.97% identity to other previously reported Per a 1 clones and 65.87-69.60% homology to the previously reported Bla g 1 clones. However, while previously reported Per a 1 clones showed homology to ANG12, a precursor protein in the midgut of the female Anopheles gambiae secreted after the blood meal, the A6 DNA sequence was found to have homology (37.1%) to DNA of G2, a putative protein in the midgut of Aedes aegypti (AY 050565). The deduced amino acid sequence of A6 contained a mitochondrial energy transfer protein signature, phosphorylation sites for the cAMP-and cGMP-dependent protein kinase C and casein kinase II. Hydrophobic and hydrophilic characteristics of the A6 deduced peptide indicated that it was a transmembrane protein. This is the first report that Per a 1 is a transmembrane protein. The deduced amino acid sequence of the A6, which contained the sequence LIRSLFGLP, differed in one amino acid from two previously reported epitopes, i.e. LIRALFGL and IRSWFGLP, of Per a 1.0104 which bound 80% and 100%, respectively, to IgE of the allergic patients tested. The A6 DNA sequence was deposited in the GenBank (Accession number AY 259514) and has been designated Per a 1.0105. The A6 expressed protein bound to monoclonal antibodies (MAb 3C2) specific to American cockroach and also bound to IgE of all (100%) of the 20 allergic Thai patients.
Diraphat
Pornphan
P
Department of Microbiology and Immunology, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand.
Sookrung
Nitat
N
Chaicumpa
Wanpen
W
Pumhirun
Phanuvich
P
Vichyanond
Pakit
P
Tapchaisri
Pramuan
P
Kalambaheti
Thareerat
T
Mahakunkijchareon
Yuvadee
Y
Sakolvaree
Yuwaporn
Y
Bunnag
Chaweewan
C
D000485:Allergens / Q000276:immunology* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D003058:Cockroaches / Q000276:immunology*; D004247:DNA / Q000235:genetics / Q000276:immunology; D015723:Gene Library; D006801:Humans; D006967:Hypersensitivity / Q000276:immunology*; D007073:Immunoglobulin E / Q000276:immunology*; D008969:Molecular Sequence Data; D013785:Thailand
D000485:Allergens; D000911:Antibodies, Monoclonal; D007073:Immunoglobulin E; D004247:DNA