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Selection of Transporter-Targeted Inhibitory Nanobodies by Solid-Supported-Membrane (SSM)-Based Electrophysiology.

Authors
  • Bärland, Natalie1
  • Perez, Camilo2
  • 1 Biozentrum, University of Basel.
  • 2 Biozentrum, University of Basel; [email protected]
Type
Published Article
Journal
Journal of Visualized Experiments
Publisher
MyJoVE Corporation
Publication Date
May 03, 2021
Issue
171
Identifiers
DOI: 10.3791/62578
PMID: 33999018
Source
Medline
Language
English
License
Unknown

Abstract

Single domain antibodies (nanobodies) have been extensively used in mechanistic and structural studies of proteins and they pose an enormous potential as tools for developing clinical therapies, many of which depend on the inhibition of membrane proteins such as transporters. However, most of the methods used to determine the inhibition of transport activity are difficult to perform in high-throughput routines and depend on labeled substrates availability thereby complicating the screening of large nanobody libraries. Solid-supported membrane (SSM) electrophysiology is a high-throughput method, used for characterizing electrogenic transporters and measuring their transport kinetics and inhibition. Here we show the implementation of SSM-based electrophysiology to select inhibitory and non-inhibitory nanobodies targeting an electrogenic secondary transporter and to calculate nanobodies inhibitory constants. This technique may be especially useful for selecting inhibitory nanobodies targeting transporters for which labeled substrates are not available.

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