Journal
/
Química
/
Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells
JoVE Journal
Química
É necessária uma assinatura da JoVE para visualizar este conteúdo.  Faça login ou comece sua avaliação gratuita.
JoVE Journal Química
Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells

Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells

DOI:
10.3791/57069-v

•

14:02 min

•

April 09, 2018

•

, , ,
1Institute of Biochemistry, Faculty of Life Sciences,University of Leipzig

Capítulos

  • 00:04Título
  • 01:11Fluorescence-based Screening of Incorporation Efficiencies
  • 04:22Genetic Incorporation of ncAAs into GPCRs
  • 08:21Ultrafast Bioorthogonal Labeling of GPCRs on Live Mammalian Cells
  • 11:11Results: Incorporation of Chemical Probes into GPCRs in Mammalian Cells for Cross-linking and Imaging Studies
  • 12:52Conclusion

Summary

Tadução automática

Tadução automática

A facile fluorescence assay is presented to evaluate the efficiency of amino-acyl-tRNA-synthetase/tRNA pairs incorporating non-canonical amino-acids (ncAAs) into proteins expressed in mammalian cells. The application of ncAAs to study G-protein coupled receptors (GPCRs) is described, including photo-crosslinking mapping of binding sites and bioorthogonal GPCR labeling on live cells.

Tags

GPCRPhoto-crosslinkingBioorthogonal ChemistryNon-canonical Amino AcidsAmber Codon SuppressionFluorescence AssayMammalian CellsProtein-protein InteractionsLigand BindingLive Cell Imaging

Article

Embed

ADICIONAR À PLAYLIST

Usage Statistics

Vídeos Relacionados

check_url/pt/57069?article_type=v

Isolation and Chemical Characterization of Lipid A from Gram-negative Bacteria
check_url/pt/57069?article_type=v

A Simple and Rapid Protocol for Measuring Neutral Lipids in Algal Cells Using Fluorescence
check_url/pt/57069?article_type=v

Preparing Adherent Cells for X-ray Fluorescence Imaging by Chemical Fixation
check_url/pt/57069?article_type=v

Chemical Vapor Deposition of an Organic Magnet, Vanadium Tetracyanoethylene
check_url/pt/57069?article_type=v

Sequencing of Plant Wall Heteroxylans Using Enzymic, Chemical (Methylation) and Physical (Mass Spectrometry, Nuclear Magnetic Resonance) Techniques
check_url/pt/57069?article_type=v

Immobilization of Multi-biocatalysts in Alginate Beads for Cofactor Regeneration and Improved Reusability
check_url/pt/57069?article_type=v

Synthesis of Protein Bioconjugates via Cysteine-maleimide Chemistry
check_url/pt/57069?article_type=v

The Preparation and Properties of Thermo-reversibly Cross-linked Rubber Via Diels-Alder Chemistry
check_url/pt/57069?article_type=v

Insights into the Interactions of Amino Acids and Peptides with Inorganic Materials Using Single-Molecule Force Spectroscopy
check_url/pt/57069?article_type=v

Preparation and Evaluation of 99mTc-labeled Tridentate Chelates for Pre-targeting Using Bioorthogonal Chemistry

Read Article