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Biotinylation of Membrane Proteins for Binder Selections


Kuhn, Benedikt T; Zimmermann, Iwan; Egloff, Pascal; Hürlimann, Lea M; Hutter, Cedric A J; Miscenic, Christian; Dawson, Roger J P; Seeger, Markus A; Geertsma, Eric R (2020). Biotinylation of Membrane Proteins for Binder Selections. In: Perez, Camilo; Maier, Timm. Expression, Purification, and Structural Biology of Membrane Proteins. Switzerland: Springer, 151-165.

Abstract

The selective immobilization of proteins represents an essential step in the selection of binding proteins such as antibodies. The immobilization strategy determines how the target protein is presented to the binders and thereby directly affects the experimental outcome. This poses specific challenges for membrane proteins due to their inherent lack of stability and limited exposed hydrophilic surfaces. Here we detail methodologies for the selective immobilization of membrane proteins based on the strong biotin-avidin interaction and with a specific focus on its application for the selection of nanobodies and sybodies. We discuss the challenges in generating and benefits of obtaining an equimolar biotin to target-protein ratio.

Abstract

The selective immobilization of proteins represents an essential step in the selection of binding proteins such as antibodies. The immobilization strategy determines how the target protein is presented to the binders and thereby directly affects the experimental outcome. This poses specific challenges for membrane proteins due to their inherent lack of stability and limited exposed hydrophilic surfaces. Here we detail methodologies for the selective immobilization of membrane proteins based on the strong biotin-avidin interaction and with a specific focus on its application for the selection of nanobodies and sybodies. We discuss the challenges in generating and benefits of obtaining an equimolar biotin to target-protein ratio.

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Additional indexing

Item Type:Book Section, refereed, original work
Communities & Collections:04 Faculty of Medicine > Institute of Medical Microbiology
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Life Sciences > Molecular Biology
Life Sciences > Genetics
Language:English
Date:2020
Deposited On:19 Jan 2021 14:36
Last Modified:20 Jan 2021 21:01
Publisher:Springer
Series Name:Methods in Molecular Biology
ISSN:1064-3745
OA Status:Hybrid
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1007/978-1-0716-0373-4_11
PubMed ID:32112321

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