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Fluorescence polarization study of a salt bridge between a single-chain Fv and its antigen ribonuclease A
- Source :
- Molecular Immunology. 34:731-734
- Publication Year :
- 1997
- Publisher :
- Elsevier BV, 1997.
-
Abstract
- The interaction between a single-chain Fv (sFv) of the monoclonal antibody 3A21 and its antigen, bovine pancreatic ribonuclease A (RNase A), was studied by site-directed mutagenesis of the hypervariable regions and fluorescence polarization analysis. The affinity constants of wild-type sFv and a mutant sFv D31A (Asp31 of heavy chain was replaced by Ala) for RNase A were found to be 2.7 x 10(7) and 4.7 x 10(6) M-1 in PBS at pH 7.2 and 37 degrees C, respectively. While the affinity constant of D31A is not affected by NaCl concentration, that of wild-type sFv is almost the same as that of D31A in the presence of more than 1 M NaCl. These results demonstrate that Asp31 of the heavy chain interacts electrostatically with a positively charged amino acid residue of RNase A.
- Subjects :
- RNase P
viruses
Immunology
Mutant
Sodium Chloride
Bovine pancreatic ribonuclease
Fluorescence Polarization Immunoassay
Asparagine
Ribonuclease
Immunoglobulin Fragments
Molecular Biology
Alanine
biology
Chemistry
Antibodies, Monoclonal
Ribonuclease, Pancreatic
Hydrogen-Ion Concentration
Molecular biology
Amino Acid Substitution
Mutagenesis, Site-Directed
biology.protein
Salts
Salt bridge
Fluorescence anisotropy
Protein Binding
Subjects
Details
- ISSN :
- 01615890
- Volume :
- 34
- Database :
- OpenAIRE
- Journal :
- Molecular Immunology
- Accession number :
- edsair.doi.dedup.....cc2491c7d109825e1de9a4947af52821