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Design of interchain disulfide bonds in the framework region of the Fv fragment of the monoclonal antibody B3
Author(s) -
Jung SunHee,
Pastan Ira,
Lee B.
Publication year - 1994
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.340190106
Subject(s) - monoclonal antibody , linker , chemistry , recombinant dna , immunoglobulin light chain , pseudomonas exotoxin , disulfide bond , fusion protein , antibody , enediyne , immunotoxin , stereochemistry , microbiology and biotechnology , computational biology , biochemistry , biology , genetics , computer science , gene , operating system
Abstract The Fv fragments are the smallest units of antibodies that retain the specific antigen binding characteristics of the whole molecule and are being used for the diagnosis and therapy of human diseases. These are noncovalently associated heterodimers of the heavy (V H ) and the light (V L ) chain variable domains, which, without modification, tend to dissociate, unfold, and/or nonspecific ally aggregate. The fragment is usually stabilized by producing it as a single chain recombinant molecule in which the two chains are linked by means of a short polypeptide linker. An alternative strategy is to connect the two chains by means of an interchain disulfide bond. We used molecular graphics and other modeling tools to identify two possible interchain disulfide bond sites in the framework region of the Fv fragment of the monoclonal mouse antibody (mAb) B3. The mAb B3 binds to many human cancer cells and is being used in the development of a new anticancer agent. The two sites identified are V H 44–V L 105 and V H 111–V L 48. (V H 44–V L 100 and V H 105–V L 43 in the numbering scheme of Kabat et al., “Sequence of Proteins of Immunological Interest,” U.S. DHHS, NIH publication No. 91–3242, 1991.) This design was recently tested using the chimeric protein composed of a truncated form of Pseudomonas exotoxin and the Fv fragment of mAb B3 with the engineered disulfide bond at V H 44–V L 105 (Brinkmann et al., Proc. Natl. Acad. Sci. U.S.A. 90:7538, 1993). The chimeric toxin was found to be just as active as the corresponding single chain counterpart and considerably more stable. Because these disulfide bond sites are in the framework region, they can be located from sequence alignment alone. We expect that the disulfide bond at these sites will stabilize the Fv fragment of most antibodies and the antigen‐specific portion of the T‐cell receptors, which are homologous. © 1994 Wiley‐Liss, Inc.