Fluorescence polarization study of a salt bridge between a single-chain Fv and its antigen ribonuclease A.

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.

journal_name

Mol Immunol

journal_title

Molecular immunology

authors

Katakura Y,Kumamoto T,Iwai Y,Kurokawa Y,Omasa T,Suga K

doi

10.1016/s0161-5890(97)00094-1

subject

Has Abstract

pub_date

1997-07-01 00:00:00

pages

731-4

issue

10

eissn

0161-5890

issn

1872-9142

pii

S0161589097000941

journal_volume

34

pub_type

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