Y55 and D78 are crucial amino acid residues of a new IgE epitope on trichosanthin.
Trichosanthin (TCS) possesses many biological and pharmaceutical activities, but its strong immunogenicity limits its clinical application. To reduce the immunogenicity of TCS, we modified the reported method for the prediction of antigenic site and identified two crucial amino acid residues (Y55 and D78) for a new epitope. We mutated these two residues into glycine and serine, respectively, and obtained three mutants, Y55G, D78S, and Y55G/D78S. These mutants induced less amount of Ig and IgG antibodies in C57BL/6J mice than wild-type TCS (wTCS) (p<0.01) and almost lost the ability to induce IgE antibody production. The mutants stimulated fewer TCS-specific B cells in C57BL/6J mice than wTCS (p<0.01). Compared with wTCS, Y55G, D78S, and Y55G/D78S lost 26.9%, 17.9%, and 98.7% specific binding ability to anti-TCS monoclonal antibody TCS4E9, respectively. These mutants still retained RNA N-glycosidase activity. In conclusion, Y55 and D78 are two crucial amino acid residues of a new IgE epitope on TCS, and their mutation reduces the immunogenicity of TCS, but still retained the enzymatic activity.
The National Laboratory of Protein Engineering and Plant Genetic Engineering, College of Life Sciences, Peking University, Beijing 100871, PR China.
Xin-Yue
XY
Zhang
Ye
Y
Wu
Jin-Yuan
JY
Yan
Yin
Y
Gao
Ye
Y
Wang
Shuang-Li
SL
Mi
Cheng-Cai
CC
An
D019943:Amino Acid Substitution; D000596:Amino Acids / Q000235:genetics / Q000276:immunology; D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology; D000939:Epitopes; D007073:Immunoglobulin E / Q000097:blood / Q000276:immunology*; D007074:Immunoglobulin G / Q000097:blood; D007136:Immunoglobulins / Q000097:blood*; D008213:Lymphocyte Activation; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009699:N-Glycosyl Hydrolases / Q000378:metabolism; D054788:Ribosome Inactivating Proteins; D015978:Trichosanthin / Q000235:genetics / Q000276:immunology*
D000596:Amino Acids; D000939:Epitopes; D007074:Immunoglobulin G; D007136:Immunoglobulins; D007073:Immunoglobulin E; D015978:Trichosanthin; D009699:N-Glycosyl Hydrolases; D054788:Ribosome Inactivating Proteins