CLONING OF COCKROACH ALLERGEN, BLA-G-4, IDENTIFIES LIGAND-BINDING PROTEINS (OR CALYCINS) AS A CAUSE OF IGE ANTIBODY-RESPONSES

An allergen cloned from a Blattella germanica (German cockroach) cDNA library, encoded a 182-amino acid protein of 20,904 Da. This protein, designated B. germanica allergen 4 (Bla g 4), was expressed as a gluta thione S-transferase fusion protein in Escherichia coil and purified b y affinity chromatography and high-performance liquid chromatography. The prevalence of serum IgE antibody to recombinant Bla g 4 in 73 cock roach allergic patients with asthma ranged from 40% (antigen binding r adioimmunoassay) to 60% (plaque immunoassay). Cockroach allergic patie nts gave positive intradermal skin tests to recombinant Bla g 4 at con centrations of 10(-3)-10(-5) mu g/ml, whereas non-allergic controls, o r cockroach allergic patients with no detectable serum IgE antibody to Bla g 4, gave negative skin tests to 1 mu g/ml. Polymerase chain reac tion and Southern analysis identified a 523-base pair DNA encoding Bla g 4 in both B. germanica and Periplaneta americana (American cockroac h). However, Northern analysis showed that mRNA encoding Bla g 4 was t ranscribed in B. germanica but not in P. americana, suggesting that al lergen expression was species specific. Sequence similarity searches s howed that Bla g 4 was a ligand binding protein or calycin and unexpec tedly revealed that this family contained several important allergens: beta-lactoglobulin, from cow milk, and rat and mouse urinary proteins . Although the overall sequence homology between these proteins was lo w (similar to 20%), macromolecular modeling techniques were used to ge nerate two models of the tertiary structure of Bla g 4, based on compa risons with the x-ray crystal coordinates of bilin binding protein and rodent urinary proteins. The results show that members of the calycin protein family can cause IgE antibody responses by inhalation or inge stion and are associated with asthma and food hypersensitivity.