Crystal structure of a dimerized cockroach allergen Bla g 2 complexed with a monoclonal antibody

The crystal structure of a 1: 1complex between the German cockroach allergen Bla g 2 and the Fab' fragment of a monoclonal antibody 7C11 was solved at 2.8-angstrom resolution. Bla g 2 binds to the antibody through four loops that include residues 60 -70, 83 -86, 98 -100, and 129 -132. Cation-Pi interactions exist between Lys65, Arg-83, and Lys-132 in Bla g 2 and several tyrosines in 7C11. In the complex with Fab ', Bla g 2 forms a dimer, which is stabilized by a quasi-four-helix bundle comprised of an alpha-helix and a helical turn from each allergen monomer, exhibiting a novel dimerization mode for an aspartic protease. A disulfide bridge between C51a and C113, unique to the aspartic protease family, connects the two helical elements within each Bla g 2 monomer, thus facilitating formation of the bundle. Mutation of these cysteines, as well as the residues Asn-52, Gln-110, and Ile-114, involved in hydrophobic interactions within the bundle, resulted in a protein that did not dimerize. The mutant proteins induced less beta hexosaminidase release from mast cells than the wild-type Bla g 2, suggesting a functional role of dimerization in allergenicity. Because 7C11 shares a binding epitope with IgE, the information gained by analysis of the crystal structure of its complex provided guidance for site-directed mutagenesis of the allergen epitope. We have now identified key residues involved in IgE antibody binding, this information will be useful for the design of vaccines for immunotherapy.

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