A bifunctional fusion protein containing Fc-binding fragment B of staphylococcal protein A amino-terminal to antidigoxin single-chain Fv
| Autor: | H. Oppermann, D. Levinson, James S. Huston, Gay-May Wu, M. Mudgett-Hunter, M S Tai, Edgar Haber |
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| Rok vydání: | 1990 |
| Předmět: |
Digoxin
Protein Conformation Recombinant Fusion Proteins viruses Immunoglobulin Fc Molecular Sequence Data Antibody Affinity Protein Engineering Biochemistry Immunoglobulin Fab Fragments Genes Synthetic Amino Acid Sequence Staphylococcal Protein A Gel electrophoresis Base Sequence biology Effector Nucleic acid sequence Antibodies Monoclonal Fusion protein Molecular biology Peptide Fragments Immunoglobulin Fc Fragments biology.protein Immunoglobulin superfamily Binding Sites Antibody Antibody Protein A Protein Binding |
| Zdroj: | Biochemistry. 29:8024-8030 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/bi00487a005 |
| Popis: | A bifunctional molecule was genetically engineered which contained an amino-terminal effector domain that bound immunoglobulin Fc (fragment B of staphylococcal protein A) and a carboxyl-terminal domain that bound digoxin [a single-chain Fv (sFv)]. Effector and sFv binding properties were virtually identical with those of the parent molecules, despite the proximity of the FB to the sFv combining site. This finding is unprecedented since in all molecules of the natural immunoglobulin superfamily, the antigen binding domain is amino terminal to the effector domain. The FB-sFv sequence was encoded in a single synthetic gene and expressed as a 33,106 molecular weight protein in Escherichia coli. After purification, renaturation, and affinity isolation, yield of active fusion protein were 110 mg/L of fermented cells (18.5-g cell paste). Bifunctionality was confirmed by the ability of FB-sFv to cross-link IgG to digoxin-bovine serum albumin, as measured by plate assays and by Ouchterlony analysis. Analysis of the expressed fusion protein suggests that the sFv holds promise for the development of multifunctional, targetable single-chain proteins. |
| Databáze: | OpenAIRE |
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