PCB-based cloning, isolation, and IgE-binding properties of recombinant latex profilin (rHev b 8)

Background: Profilin(Hev b 8) in natural rubber latex (NRL) has been assume d to be an important allergen. Since latex profilin has a molecular mass si milar to two other latex allergens (Hev b 1 and Hev b 6.03) in the 14-kDa r ange, it is difficult to obtain sufficient amounts of purified native profi lin for investigations and diagnostics. The present study aimed to produce recombinant latex profilin (rHev b 8) and study its IgE-binding reactivity. Methods: A profilin-specific cDNA encoding the latex profilin from Hevea br asiliensis leaves was synthesized and subcloned, and the rHev b 8 was overe xpressed in fusion with the maltose-binding protein (MBP) in E. coli. The I gE-binding reactivity of rHev b 8 was studied by immunoblotting, immunoblot inhibition experiments, and the Pharmacia CAP method, with 25 sera from he alth-care workers with latex allergy and 17 sera from latex-sensitive spina bifida patients. Results: rHev b 8 was found to have 131 amino acids and a sequence identity of 75% with birch profilin (Bet v 2). Analysis by the CAP system revealed the presence of rHev b 8-specific IgE antibodies in two out of 17 sera from spina bifida patients and in five out of 25 sera (20%) from health-care wo rkers. Two subjects of the latter group with rHev b 8-specific IgE showed n egative results in the skin prick tests with tree-pollen extracts and had n o IgE to rBet v 2, indicating the presence of IgE-binding epitopes on the H ev b 8-molecule which do not cross-react with birch profilin. Immunoblot in hibition assays using MBP-rHev b 8 as inhibitor confirmed the presence of l atex profilin in the NRL extract. IgE binding to the native latex profilin could be completely inhibited by the MBP-rHev b 8. Conclusions: Latex profilin represents a minor allergen in NRL and may have IgE-binding epitopes different from Bet v 2.