Identification of IgE binding epitopes of peptide fragments of Bla g 6 from German cockroach (Blattella germanica)

Abstract

German cockroach allergens induce IgE-mediated hypersensitivity. Among the German cockroach allergens studied, Bla g 6 shows homology to muscle protein troponin C. It contains four calcium-binding domains at 20-30, 56-67, 96-107, and 132-143 amino acid (aa) residues, and its immunoglobulin E (IgE) reactivity is dependent upon calcium ion level. However, the IgE binding epitopes of Bla g 6 have not been studied. This study aimed to identify the IgE binding epitopes from the five linear peptide fragments of Bla g 6. The full-length and five peptide fragments (P1: aa 1-111, P2: aa 1-95, P3: aa 33-111, P4: aa 80-151, and P5: aa 33-151) of Bla g 6 were generated by polymerase chain reaction (PCR) and expressed in Escherichia coli (E. coli). Enzyme-linked immunosorbent assay (ELISA) was performed on 24 patients’ sera that adjusted the final concentration 10 mM of CaCl2 to determine the IgE activities of Bla g 6. Eight sera (33.3%), 9 sera (37.5%), and 11 sera (45.8%) showed IgE reactivity to Bla g 6.0101, Bla g 6.0201, and Bla g 6.0301, respectively. Among the sera from the positive IgE reactivity, three patients’ sera were selected and the IgE reactivity was measured by ELISA with the five recombinant peptide fragments of Bla g 6. Based on IgE responses, one patient’s serum exhibited the strongest IgE reactivity. I assumed that the aa between 96-151 residues, including the calcium binding domains III and IV, would be important for IgE binding. These results may provide information that will yield safe diagnostic methods and immunotherapeutics.