TY - JOUR
T1 - Analysis of the three-dimensional antigenic structure of giant ragweed allergen, Amb t 5
AU - Rafnar, Thorunn
AU - Brummet, Mary E.
AU - Bassolino-Klimas, Donna
AU - Metzler, William J.
AU - Marsh, David G.
N1 - Funding Information:
We thank Dr. David G. Klapper for providing reduced and alkylated rAmb t 5 and for technical advice. We thank Dr. Amina Woods for assistance in the purification of rAmb t 5. This work was supported by National Institutes of Health Grant AI19727.
PY - 1998/5/1
Y1 - 1998/5/1
N2 - The ragweed allergens Amb t 5 and Amb a 5 are among the smallest inhaled protein allergens known, containing a single, immunodominant T-cell epitope. In this study we analyzed the B-cell epitope structure of Amb t 5. The three- dimensional structures of Amb t 5 and Amb a 5 have been determined by NMR spectroscopy, providing a rare opportunity to analyze three-dimensional antigenic sites. Amb t 5 residues likely to be important for antigenicity were identified by examining the surface area of Amb t 5 accessible to a probe of the size of an antibody molecule. After changing these residues to the corresponding Amb a 5 residues, recombinant proteins were purified and tested for loss of antigenic activity. Inhibition radio-immunoassays, using sera from 8 individuals who had received immunotherapy with giant ragweed extract, allowed the mutations to be divided into three groups: (1). mutations that had little or no effect on antibody binding, (2) mutations that caused a loss of antigenic activity to a different degree in different sera and (3) mutations that drastically reduced antigenic activity in all sera tested. This last set of mutations clustered in the third loop of Amb t 5, suggesting that antibody recognition of Amb t 5, like T-cell recognition, is primarily directed towards a single, immunodominant site.
AB - The ragweed allergens Amb t 5 and Amb a 5 are among the smallest inhaled protein allergens known, containing a single, immunodominant T-cell epitope. In this study we analyzed the B-cell epitope structure of Amb t 5. The three- dimensional structures of Amb t 5 and Amb a 5 have been determined by NMR spectroscopy, providing a rare opportunity to analyze three-dimensional antigenic sites. Amb t 5 residues likely to be important for antigenicity were identified by examining the surface area of Amb t 5 accessible to a probe of the size of an antibody molecule. After changing these residues to the corresponding Amb a 5 residues, recombinant proteins were purified and tested for loss of antigenic activity. Inhibition radio-immunoassays, using sera from 8 individuals who had received immunotherapy with giant ragweed extract, allowed the mutations to be divided into three groups: (1). mutations that had little or no effect on antibody binding, (2) mutations that caused a loss of antigenic activity to a different degree in different sera and (3) mutations that drastically reduced antigenic activity in all sera tested. This last set of mutations clustered in the third loop of Amb t 5, suggesting that antibody recognition of Amb t 5, like T-cell recognition, is primarily directed towards a single, immunodominant site.
KW - Allergens
KW - Epitope structure
KW - Ragweed
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U2 - 10.1016/S0161-5890(99)00054-1
DO - 10.1016/S0161-5890(99)00054-1
M3 - Article
C2 - 9798650
AN - SCOPUS:0031692206
SN - 0161-5890
VL - 35
SP - 459
EP - 467
JO - Molecular Immunology
JF - Molecular Immunology
IS - 8
ER -