A Structure-Based Database of Antibody Variable Domain Diversity

The diversity of natural antibodies is limited by the genetic mechanisms that engender diversity and the functional requirements of antigen binding. Using an in vitro-evolved autonomous heavy chain variable domain (V HH-RIG), we have investigated the limits of structurally-tolerated diversity in the...

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Bibliographic Details
Published in:Journal of molecular biology 2005-05, Vol.348 (3), p.699-709
Main Authors: Bond, Christopher J., Wiesmann, Christian, Marsters, James C., Sidhu, Sachdev S.
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
antibody
Antibody Diversity
combinatorial mutagenesis
Complementarity Determining Regions - chemistry
Complementarity Determining Regions - genetics
Crystallography, X-Ray
Humans
Immunoglobulin Heavy Chains - chemistry
Immunoglobulin Heavy Chains - genetics
Models, Molecular
Molecular Sequence Data
Peptide Library
phage display
protein engineering
Protein Structure, Tertiary
variable domain
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Summary:The diversity of natural antibodies is limited by the genetic mechanisms that engender diversity and the functional requirements of antigen binding. Using an in vitro-evolved autonomous heavy chain variable domain (V HH-RIG), we have investigated the limits of structurally-tolerated diversity in the three complementarity-determining regions and a fourth loop within the third framework region. We determined the X-ray crystal structure of the V HH-RIG domain at 1.9 Å resolution and used it to guide the design of phage-displayed libraries encompassing the four loops. The libraries were subjected to selections for structural stability, and a database of structurally-tolerated sequences was compiled from the sequences of approximately 1000 unique clones. The results reveal that all four loops accommodate significantly greater diversity than is observed in nature. Thus, it appears that most sequence biases in the natural immune repertoire arise from factors other than structural constraints and, consequently, it should be possible to enhance the functions of antibodies significantly through in vitro evolution.
ISSN:0022-2836
1089-8638
DOI:10.1016/j.jmb.2005.02.063