Molecular and idiotypic analysis of antibodies to Cryptococcus neoformans glucuronoxylomannan

Antibodies to the Cryptococcus neoformans capsular glucuronoxylomannan (GXM) form the basis of two potential therapeutic intervention strategies, i.e., conjugate vaccines and passive antibody therapy. To better understand the molecular basis of the antibody response, the heavy- and light-chain immunoglobulin variable region (V(H) and V(L), respectively) sequences of seven monoclonal antibodies (MAbs) to GXM were determined. Rabbit anti- idiotypic serum was made to the previously characterized murine MAb 2H1 and used to study MAb 2H1 idiotype expression in other GXM-binding MAbs and immune sera. MAb E1 originated from a C3H/HeJ mouse immunized with C. neoformans serotype A polysaccharide. MAbs 471, 1255, 339, 3C2, 386, and 302 originated from BALB/c mice immunized with polysaccharide of serotypes A, A, B, C, D, and D, respectively, conjugated to sheep erythrocytes. In the E1, V(H) uses V(H) from the T15 gene family and J(H)3 and has a D segment of three amino acids, and the V(L) uses a V(κ)Ser-like gene family element and J(κ)5. In MAbs 471 and 3C2, the V(H) uses V(H)7183-like gene family elements and J(H)2 and has D segments of seven amino acids, and the V(L) uses V(κ)5.1 and J(κ)1. In MAbs 1255 and 339, the V(H) uses V(H)10-like gene elements and J(H)4 and has six codon D segments, and the V(L) uses a V(κ)21-like gene element and J(κ)5. In MAbs 302 and 386, respectively, the V(H) uses V(H)GAM- like gene elements and J(H)2 and J(H)3 and has six and four codon D segments, and V(L) uses V(κ)4/5-like gene elements and J(κ)1. V(H) usage, MAb 2H1 idiotype expression, and fine specificity mapping define a minimum of three GXM epitopes which elicit protective antibodies. The results confirm that the antibody response is highly restricted, suggest a close relationship between molecular structure and serological properties, and provide insight into protein structural motifs important for GXM binding.