Combining Conformational Profiling of GPCRs with CRISPR/Cas9 Gene Editing Approaches

Kyla Bourque, Dominic Devost, Asuka Inoue, Terence E. Hébert

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Ligand-biased signaling could have a significant impact on drug discovery programs. As such, many approaches to screening now target a larger section of the signaling responses downstream of an individual G protein-coupled receptor (GPCR). Biosensor-based platforms have been developed to capture signaling signatures. Despite the ability to use such signaling signatures, they may still be particular to an individual cell type and thus such platforms may not be portable from cell to cell, necessitating further cell-specific biosensor development. We have developed a complementary strategy based on capturing receptor-proximal conformational profiles using intra-molecular BRET-based sensors composed of a Renilla luciferase donor engineered into the carboxy-terminus and CCPGCC motifs which bind fluorescent hairpin biarsenical dyes engineered into different positions into the receptor primary structure. Here, we discuss how these experiments can be conducted and combined with CRISPR/Cas9 genome editing to assess specific G protein-dependent and -independent events.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages169-182
Number of pages14
DOIs
Publication statusPublished - 2019 Jan 1

Publication series

NameMethods in Molecular Biology
Volume1947
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • BRET
  • CRISPR
  • Conformation-sensitive biosensors
  • Drug discovery
  • GPCRs
  • Genome-editing

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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  • Cite this

    Bourque, K., Devost, D., Inoue, A., & Hébert, T. E. (2019). Combining Conformational Profiling of GPCRs with CRISPR/Cas9 Gene Editing Approaches. In Methods in Molecular Biology (pp. 169-182). (Methods in Molecular Biology; Vol. 1947). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-9121-1_9