Analysis of the three-dimensional antigenic structure of giant ragweed allergen, Amb t 5.
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.
Rafnar
T
T
Division of Clinical Immunology, Johns Hopkins Asthma and Allergy Center, Johns Hopkins University School of Medicine, Baltimore, MD 21224, USA. rafnar@krabb.is
Brummet
M E
ME
Bassolino-Klimas
D
D
Metzler
W J
WJ
Marsh
D G
DG
D000485:Allergens / Q000737:chemistry*; D000596:Amino Acids / Q000235:genetics; D000917:Antibody Formation; D000937:Antigen-Antibody Reactions / Q000235:genetics; D052179:Antigens, Plant; D001666:Binding Sites, Antibody / Q000235:genetics; D006801:Humans; D016296:Mutagenesis; D009154:Mutation / Q000502:physiology; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D011058:Pollen / Q000276:immunology*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics
D000485:Allergens; D000596:Amino Acids; D052179:Antigens, Plant; D010940:Plant Proteins; D011994:Recombinant Proteins; C069543:group 5 allergens, Ambrosia