Genetic code expansion to enable site-specific bioorthogonal labeling of functional G protein-coupled receptors in live cells.

ABSTRACT: For use in site-specific bioorthogonal labeling of expressed G protein-coupled receptors (GPCRs) in live cells, we developed a luciferase-based reporter assay. The assay was used to compare amber codon suppression efficiency, receptor functionality, and efficiency of different bioorthogonal labeling chemistries. We used the assay system to compare side-by-side the efficiency of incorporation of three different noncanonical amino acids [4-azido-l-phenylalanine (azF), cyclopropene-l-lysine (CpK), and trans-cyclooct-2-en-l-lysine (TCOK)] at three different sites on a GPCR using three different genetic code expansion plasmid systems. As a model GPCR, we engineered an epitope-tagged C-C chemokine receptor 5 (CCR5)-RLuc3 fusion for expression in HEK293T cells. Satisfactory incorporation of azF,

SUBMITTER: Mattheisen JM 

PROVIDER: S-EPMC9847076 | biostudies-literature | 2023 Feb

REPOSITORIES: biostudies-literature