Elimination of a human T-cell region in staphylokinase by T-cell screening and computer modeling.
Staphylokinase is a potent highly fibrin-selective thrombolytic agent, but it induces a humoral immune response in most treated patients. Staphylokinase-specific T-lymphocytes can be found in normal healthy individuals, from whom a large panel of staphylokinase-specific T-cells were cloned. The staphylokinase amino acid sequence 71-87 was widely recognized, as it induced proliferation of T-cell clones isolated from 90% of the donors. Computer modeling of this area, threaded as 11-mer peptides within the peptide-binding groove of the major HLA-DR alleles, indicated two putative partially overlapping binding sequences. The region-(71-87)-specific T-cell clones recognized either one or the other minimal peptide, confirming that both sequences could be functional T-cell epitopes. Furthermore, to guide the mutagenesis to eliminate T-cell reactivity, the contribution of each residue to the HLA-DR-anchoring and T-cell receptor exposure was evaluated for both binding motifs. Computer calculations combined with functional assays resulted in the design of staphylokinase-variants, including 2 to 4 amino acid substitutions in the region 71-87. These variants were no longer recognized by the region-(71-87)-specific T-cell clones, and importantly no new staphylokinase-variant-specific cellular immune response could be measured.
Warmerdam
Petra A M
PA
Center for Transgene Technology and Gene Therapy, Flanders Interuniversity Institute for Biotechnology, Leuven, Belgium. petra.warmerdam@med.kuleuven.ac.be
Plaisance
Stéphane
S
Vanderlick
Kristel
K
Vandervoort
Petra
P
Brepoels
Kathleen
K
Collen
Désiré
D
De Maeyer
Marc
M
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001665:Binding Sites; D003198:Computer Simulation; D015195:Drug Design; D000939:Epitopes / Q000737:chemistry / Q000276:immunology; D005343:Fibrinolytic Agents / Q000737:chemistry / Q000276:immunology*; D006684:HLA-DR Antigens / Q000276:immunology; D006801:Humans; D008213:Lymphocyte Activation; D008666:Metalloendopeptidases / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009842:Oligopeptides / Q000737:chemistry / Q000276:immunology; D011487:Protein Conformation; D016176:T-Lymphocyte Subsets / Q000276:immunology*
D000942:Antigens, Bacterial; D000939:Epitopes; D005343:Fibrinolytic Agents; D006684:HLA-DR Antigens; D009842:Oligopeptides; D008666:Metalloendopeptidases; C022653:auR protein, Staphylococcus aureus