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Engineered peptide barcodes for in-depth analyses of binding protein libraries


Egloff, Pascal; Zimmermann, Iwan; Arnold, Fabian M; Hutter, Cedric A J; Morger, Damien; Opitz, Lennart; Poveda, Lucy; Keserue, Hans-Anton; Panse, Christian; Roschitzki, Bernd; Seeger, Markus A (2019). Engineered peptide barcodes for in-depth analyses of binding protein libraries. Nature Methods, 16(5):421-428.

Abstract

Binding protein generation typically relies on laborious screening cascades that process candidate molecules individually. We have developed NestLink, a binder selection and identification technology able to biophysically characterize thousands of library members at once without the need to handle individual clones at any stage of the process. NestLink uses genetically encoded barcoding peptides termed flycodes, which were designed for maximal detectability by mass spectrometry and support accurate deep sequencing. We demonstrate NestLink's capacity to overcome the current limitations of binder-generation methods in three applications. First, we show that hundreds of binder candidates can be simultaneously ranked according to kinetic parameters. Next, we demonstrate deep mining of a nanobody immune repertoire for membrane protein binders, carried out entirely in solution without target immobilization. Finally, we identify rare binders against an integral membrane protein directly in the cellular environment of a human pathogen. NestLink opens avenues for the selection of tailored binder characteristics directly in tissues or in living organisms.

Abstract

Binding protein generation typically relies on laborious screening cascades that process candidate molecules individually. We have developed NestLink, a binder selection and identification technology able to biophysically characterize thousands of library members at once without the need to handle individual clones at any stage of the process. NestLink uses genetically encoded barcoding peptides termed flycodes, which were designed for maximal detectability by mass spectrometry and support accurate deep sequencing. We demonstrate NestLink's capacity to overcome the current limitations of binder-generation methods in three applications. First, we show that hundreds of binder candidates can be simultaneously ranked according to kinetic parameters. Next, we demonstrate deep mining of a nanobody immune repertoire for membrane protein binders, carried out entirely in solution without target immobilization. Finally, we identify rare binders against an integral membrane protein directly in the cellular environment of a human pathogen. NestLink opens avenues for the selection of tailored binder characteristics directly in tissues or in living organisms.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Functional Genomics Center Zurich
04 Faculty of Medicine > Institute of Medical Microbiology
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Life Sciences > Biotechnology
Life Sciences > Biochemistry
Life Sciences > Molecular Biology
Life Sciences > Cell Biology
Language:English
Date:May 2019
Deposited On:06 Jan 2020 13:28
Last Modified:29 Jul 2020 12:05
Publisher:Nature Publishing Group
ISSN:1548-7091
OA Status:Closed
Publisher DOI:https://doi.org/10.1038/s41592-019-0389-8
PubMed ID:31011184

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