Advertisement

Incorporation of the Unnatural Amino Acid p-benzoyl-L-phenylalanine (Bpa) into a G Protein-coupled Receptor in its Native Context

  • George Umanah
  • Li-Yin Huang
  • Peter G. Schultz
  • Fred Naider
  • Jeffrey M. Becker
Part of the Advances in Experimental Medicine and Biology book series (volume 611)

Introduction

Ligand binding initiates a change in the conformation of G protein-coupled receptors (GPCRs) resulting in activation of the G protein-mediated signal transduction cascade [1]. We are studying a novel approach to elucidate the dynamics of GPCR structure by the co-translational introduction of unnatural amino acids (UAAs) into the receptor.

UAAs can be synthesized to contain a variety of chemical moieties for use as photoaffinity labels, fluorescent labels, or spectroscopic probes. Orthogonal tRNA/ aminoacyl-tRNA synthetase pairs evolved and expressed in the target cell have been used to incorporate UAAs into heterologously expressed protein in living cells [2]. The mutated tRNA, charged with its UAA, recognizes the amber TAG stop codon and incorporates the non-natural amino acid into the nascent polypeptide chain (See Figure below). UAAR has been widely used in the heterologous Xenopusoocyte expression system to insert UAAs into a variety of receptors and channel...

Keywords

Unnatural Amino Acid Aminoacyl tRNA Synthetase Amino Acid Analog Nascent Polypeptide Chain Eukaryotic Host Cell 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

This work was supported by grants NIH GM-22086(FN) and NIH GM-22087(JMB).

References

  1. 1.
    Eilers, M., et al. Biochemistry 44, 8959–8975 (2005).CrossRefGoogle Scholar
  2. 2.
    Wang, L., Xie, J., and Schultz, P. G. Annu. Rev. Biophys. Biomol. Struct. 35, 225–249 (2006).CrossRefGoogle Scholar
  3. 3.
    Williams, I., et al. Nucleic Acids Res 32, 6605–6616 (2004).CrossRefGoogle Scholar
  4. 4.
    Lee, B. K., et al. Biochemistry 41, 13681–13689 (2002)CrossRefGoogle Scholar
  5. 5.
    Island, M. D., et al. Curr. Genet. 20, 457–463 (1991).CrossRefGoogle Scholar
  6. 6.
    Hauser, M., et al. Mol. Membr. Biol. 22, 215–27 (2005).CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • George Umanah
    • 1
  • Li-Yin Huang
    • 1
  • Peter G. Schultz
    • 2
  • Fred Naider
    • 3
  • Jeffrey M. Becker
    • 1
  1. 1.Department of MicrobiologyUniversity of TennesseeKnoxville
  2. 2.Department of ChemistryThe Scripps Research InstituteLa Jolla
  3. 3.Department of ChemistryCollege of Staten IslandCUNY

Personalised recommendations