IMMUNOLOGICAL AND BIOLOGICAL PROPERTIES OF RECOMBINANT LOL-P-1

Background: Current forms of allergy diagnosis and therapies are based on the use of natural allergenic extracts. Despite strong evidence th at higher therapeutic efficacy may be achieved with purified allergens , the purification of multiple allergic components from extracts is a fastidious and sometimes an impossible task. However, the use of recom binant allergens may be an alternative to overcome this problem. Objec tive: In this study, we compared the immunological properties of recom binant (r) Lol p 1 with those of the natural protein. Method: We clone d directly the gene encoding Lol p 1 from genomic DNA of ryegrass poll en. This gene was subcloned into the expression vector pMAL(R)-c and e xpressed as fusion protein. Subsequently, rLol p 1 was cleaved from ma ltose-binding protein using factor Xa. Using binding inhibition and pr oliferative assays, we assessed the immunological properties of the re combinant allergens. The capacity of rLol p 1 to trigger basophil hist amine release and to elicit a skin reaction was also assessed and comp ared to those of its natural counterpart. Results: We found that the L ol p 1 gene has no introns since we amplified this gene directly from genomic DNA. We demonstrated that the binding sites of anti-Lol p 1 mo noclonal antibody, specific human IgG and IgE antibody are well conser ved on rLol p 1 as no difference in the binding inhibition profile was observed when using either natural or recombinant protein. At the T-c ell level, rLol p 1 elicited a T-cell response in mice comparable to t hat observed with the natural protein. In addition, we demonstrated th at the biological characteristics of rLol p 1 were comparable to those of the natural counterpart, in that rLol p 1 elicited a skin wheal re action and induced basophil histamine release in grass-allergic patien ts only. Conclusion: The data indicate that natural Lol p 1 and rLol p 1 shared identical immunological and biological properties.