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Determination of binding site of anti‐tumour necrosis factor‐α monoclonal antibody using hybrid and mutant proteins
Author(s) -
Shingarova Ludmila N.,
Sagaidak Lubov N.,
Berkova Nadia,
Korobko Vyacheslav G.
Publication year - 1996
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(96)00396-1
Subject(s) - monoclonal antibody , microbiology and biotechnology , epitope , biology , fusion protein , mutant , antibody , coding region , escherichia coli , tumor necrosis factor alpha , gene , virology , recombinant dna , genetics , immunology
In order to map the immunogenic epitope for the monoclonal antibody E7H2 on the human tumour necrosis factor (hTNF‐α) molecule, a number of chimeric proteins were developed by in‐frame joining segments of the human genes encoding TNF‐α and lymphotoxin (TNF‐β) as well as by coupling appropriate coding regions for human and mouse TNF‐α. High level expression of these chimeric genes was achieved in Escherichia coli by placing the coding sequences under control of either E. coli trp ‐promoter or a tandem of bacteriophage T7 constitutive promoters A2 and A3. As revealed by Western blot analysis with monoclonal antibody E7H2 directed against human TNF‐α, the region involved in the binding of this antibody includes sequence ValGluLeuArg in the N‐terminal part of the TNF‐α molecule.