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10 Å resolution followed by molecular modeling. Both antibodies engage residues on the protruding spikes but their epitopes and binding orientations differ. Steric interference effects limit maximum binding to 
doi:10.1016/j.jmb.2006.10.088 
Copyright © 2006 Elsevier Ltd All rights reserved.
Structural Basis of Enhanced Binding of Extended and Helically Constrained Peptide Epitopes of the Broadly Neutralizing HIV-1 Antibody 4E10
Rosa M.F. Cardoso1, Florence M. Brunel2, Sharon Ferguson1, Michael Zwick3, Dennis R. Burton1, 3, Philip E. Dawson2, 4 and Ian A. Wilson1, 4, 
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Received 31 July 2006;
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Abstract
Potent, broadly HIV-1 neutralizing antibodies (nAbs) may be invaluable for the design of an AIDS vaccine. 4E10 is the broadest HIV-1 nAb known to date and recognizes a contiguous and highly conserved helical epitope in the membrane-proximal region of gp41. The 4E10 epitope is thus an excellent target for vaccine design as it is also highly amenable to peptide engineering to enhance its helical character. To investigate the structural effect of both increasing the peptide length and of introducing helix-promoting constraints in the 4E10 epitope, we have determined crystal structures of Fab 4E10 bound to an optimized peptide epitope (NWFDITNWLWYIKKKK-NH2), an Aib-constrained peptide epitope (NWFDITNAibLWRR-NH2), and a thioether-linked peptide (NWFCITOWLWKKKK-NH2) to resolutions of 1.7 Å, 2.1 Å, and 2.2 Å, respectively. The thioether-linked peptide is the first reported structure of a cyclic tethered helical peptide bound to an antibody. The introduced helix constraints limit the conformational flexibility of the peptides without affecting interactions with 4E10. The substantial increase in affinity (10 nM versus 104 nM of the IC50 of the original KGND peptide template) is largely realized by 4E10 interaction with an additional helical turn at the peptide C terminus that includes Leu679 and Trp680. Thus, the core 4E10 epitope was extended and modified to a WFX(I/L)(T/S)XX(L/I)W motif, where X does not play a major role in 4E10 binding and can be used to introduce helical-promoting constraints in the peptide epitope.
Keywords: 4E10; gp41; crystal structure; antibody; AIDS vaccine
Abbreviations: HIV-1, human immunodeficiency virus type 1; CDR, complementarity-determining region; Aib, α-amino isobutyric acid
Article Outline
- Introduction
- Results
- Structure determination and refinement
- Structure of Fab 4E10
- Structure of the peptide epitope
- The combining site
- Expansion of the core epitope
- Discussion
- Material and Methods
- Peptide synthesis and purification
- Preparation of complexes, crystallization and data collection
- Structure determination and refinement
- Structural analysis
- Binding affinity by ELISA
- Binding affinity by surface plasmon resonance
- Protein Data Bank accession codes
- Acknowledgements
- References
