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Nat Struct Mol Biol. 2018 Mar;25(3):289-296. doi: 10.1038/s41594-018-0028-6. Epub 2018 Feb 12.

Yeast surface display platform for rapid discovery of conformationally selective nanobodies.

Author information

1
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA, USA.
2
Department of Neuroscience, University of Copenhagen, Copenhagen, Denmark.
3
Department of Molecular and Cellular Physiology, Stanford University, Stanford, CA, USA.
4
Department of Immunobiology, Yale School of Medicine, New Haven, CT, USA.
5
Department of Pharmaceutical Chemistry, University of California San Francisco, San Francisco, CA, USA. Aashish.Manglik@ucsf.edu.
6
Department of Anesthesia and Perioperative Care, University of California San Francisco, San Francisco, CA, USA. Aashish.Manglik@ucsf.edu.
7
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA, USA. Andrew_Kruse@hms.harvard.edu.

Abstract

Camelid single-domain antibody fragments ('nanobodies') provide the remarkable specificity of antibodies within a single 15-kDa immunoglobulin VHH domain. This unique feature has enabled applications ranging from use as biochemical tools to therapeutic agents. Nanobodies have emerged as especially useful tools in protein structural biology, facilitating studies of conformationally dynamic proteins such as G-protein-coupled receptors (GPCRs). Nearly all nanobodies available to date have been obtained by animal immunization, a bottleneck restricting many applications of this technology. To solve this problem, we report a fully in vitro platform for nanobody discovery based on yeast surface display. We provide a blueprint for identifying nanobodies, demonstrate the utility of the library by crystallizing a nanobody with its antigen, and most importantly, we utilize the platform to discover conformationally selective nanobodies to two distinct human GPCRs. To facilitate broad deployment of this platform, the library and associated protocols are freely available for nonprofit research.

Comment in

PMID:
29434346
PMCID:
PMC5839991
DOI:
10.1038/s41594-018-0028-6
[Indexed for MEDLINE]
Free PMC Article

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