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Edited by K. C. (Chris) Christopher Garcia, Stanford University, Stanford, CA, and approved July 10, 2019 (received for review June 16, 2019)
Significance
Antibodies have been found to be helpful for structural studies of challenging proteins by X-ray crystallography and electron microscopy. They are being used to construct useful protein nanostructures as well. Antibodies suitable for structural study should recognize the 3-dimensional conformations of target proteins. However, generating such antibodies and characterizing structures of their complexes with antigens take months or even years of research. Here, we show that we can expand the application of well-characterized antibodies by “transplanting” the alpha-helical epitopes that they recognize to proteins with completely different structures and sequences. Systematic screening of more antihelix antibodies will greatly expand the scope of the method.
Abstract
Antibodies are indispensable tools in protein engineering and structural biology. Antibodies suitable for structural studies should recognize the 3-dimensional (3D) conformations of target proteins. Generating such antibodies and characterizing their complexes with antigens take a significant amount of time and effort. Here, we show that we can expand the application of well-characterized antibodies by “transplanting” the epitopes that they recognize to proteins with completely different structures and sequences. Previously, several antibodies have been shown to recognize the alpha-helical conformation of antigenic peptides. We demonstrate that these antibodies can be made to bind to a variety of unrelated “off-target” proteins by modifying amino acids in the preexisting alpha helices of such proteins. Using X-ray crystallography, we determined the structures of the engineered protein–antibody complexes. All of the antibodies bound to the epitope-transplanted proteins, forming accurately predictable structures. Furthermore, we showed that binding of these antihelix antibodies to the engineered target proteins can modulate their catalytic activities by trapping them in selected functional states. Our method is simple and efficient, and it will have applications in protein X-ray crystallography, electron microscopy, and nanotechnology.
Footnotes
↵1J.W.K. and S.K. contributed equally to this work.
- ↵2To whom correspondence may be addressed. Email: misunjin{at}gist.ac.kr or jieoh{at}postech.ac.kr.
Author contributions: J.W.K., S.K., Haerim Lee, Hayyoung Lee, M.S.J., and J.-O.L. designed research; J.W.K., S.K., Haerim Lee, and G.C. performed research; J.W.K., S.K., Haerim Lee, G.C., and J.-O.L. analyzed data; and J.W.K., S.K., Haerim Lee, S.C.K., Hayyoung Lee, M.S.J., and J.-O.L. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
Data deposition: The atomic coordinates and X-ray diffraction data reported in this paper have been deposited in the Protein Data Bank (PDB) under the following ID codes: 6K68 for 8420-3MNZ, 6K65 for 9014-1P4B, 6K64 for 8188-3LRH, 6K3M for 8189-3LRH, 6K6B for 8496-3LRH, 6K67 for 9011-3LRH, 6K69 for 9213-3LRH, and 6K6A for 8188cys-3LRHcys.
See Commentary on page 17611.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1910080116/-/DCSupplemental.
Published under the PNAS license.
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Application of antihelix antibodies in protein structure determination
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