Simultaneous removal of T and B cell epitope in recombinant staphylokinase by structure-based mutagenesis of immuno-dominant Arg77 and Glu80 residues
Arg77和Glu80定点突变同时去除葡激酶中的T和B细胞抗原表位

Objective]To reduce immunogenicity of recombined staphylokinase(r-yak),site-directed mutagenesis of Arg77 and Glu80 residue was performed to simultaneously remove T and B cell epitope in r-Sak molecule.Methods The solvent accessible surface areas of residues 77 and 80 in r-Sak were used to analyze rational design of Sak mutation.The Sak mutants were expressed in E coli DHSa.After purified by a 3-step chromatography,their fibrinolytic activities and immunological properties were analyzed.Results]Immunogenicity tests suggested that Sak induced a Th2-type immune response.Substitution of Glu80 with alanine or serine successfully reduced its solvent accessible surface area while simultaneously removing part of the T and B cell epitope.Changing Arg77 to glutamine,asparagine,or lysine removed only part of the T cell epitope.Of six dually substituted variants,Sak(R77Q/E80A)and Sak(R77Q/E80S)variants effectively eliminated part of the B and T cell epitopes,which markedly reduced their immunogenicity.