CLONING, EXPRESSION, AND IMMUNOLOGICAL CHARACTERIZATION OF RECOMBINANT LOLIUM-PERENNE ALLERGEN LOL P-II

The molecular cloning of the cDNA encoding for an isoallergenic form o f Lol p II, a major rye grass (Lolium perenne) pollen allergen, was pe rformed by polymerase chain reaction amplification on mRNA extracted f rom pollen. The amino acid sequence derived from the cDNA was truncate d by 4 and 5 residues at the NH2- and COOH-terminal ends, respectively , and differed only in one position from that previously reported. Thi s cDNA was expressed in Escherichia coli by fusion to the carboxyl ter minus of the human ferritin H-chain. The molecule was produced in high yields as a soluble protein and was easily purified. The protein reta ins the multimeric quaternary structure of ferritin, and it exposes on the surface the allergenic moiety, which can be recognized in Western blotting and in enzyme-linked immunosorbent assay experiments by spec ific IgE from allergic patients. The recombinant allergen was used to analyze the sera of 26 patients allergic to L. perenne compared with c ontrol sera. The results were in good agreement with the values obtain ed with the radioallergosorbent test assay. In addition, histamine rel ease experiments in whole blood from an allergic patient and skin pric k tests showed that the recombinant allergen retains some of the biolo gical properties of the natural compound. These findings indicate that the availability of homogeneous recombinant allergens may be useful f or the development of more specific diagnostic and therapeutic procedu res. Moreover, this expression system may be of more general interest for producing large amounts of soluble protein domains in E. coli.