Reproducing the natural evolution of protein structural features with the selectively infective phage (SIP) technology. The kink in the first strand of antibody kappa domains.
- Spada S Biochemisches Institut der Universität Zürich, Winterthurerstrasse 190, Zürich, CH-8057, Switzerland.
- Honegger A
- Plückthun A
- 1998-10-14
Published in:
- Journal of molecular biology. - 1998
Bacteriophages
Directed Molecular Evolution
Escherichia coli
Immunoglobulin kappa-Chains
Models, Molecular
Protein Conformation
Protein Denaturation
Selection, Genetic
Urea
English
The beta-sandwich structure of immunoglobulin variable domains is characterized by a typical kink in the first strand, which allows the first part of the strand to hydrogen bond to the outer beta-sheet (away from the VH-VL interface) and the second part to the inner beta-sheet. This kink differs in length and sequence between the Vkappa, Vlambda and VH domains and yet is involved in several almost perfectly conserved interactions with framework residues. We have used the selectively infective phage (SIP) system to select the optimal kink region from several defined libraries, using an anti-hemagglutinin single-chain Fv (scFv) fragment as a model system. Both for the kink with the Vkappa domain length and that with the Vlambda length, a sequence distribution was selected that coincides remarkably well with the sequence distribution of natural antibodies. The selected scFv fragments were purified and characterized, and thermodynamic stability was found to be the prime factor responsible for selection. These data show that the SIP technology can be used for optimizing protein structural features by evolutionary approaches.
- Language
-
- English
- Open access status
- green
- Identifiers
-
- DOI 10.1006/jmbi.1998.2068
- PMID 9769213
- Persistent URL
- https://sonar.ch/global/documents/237853
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