Probing Subunits Interactions in KATP Channels Using Photo-Crosslinking via Genetically Encoded p-Azido-l-phenylalanine

Prasanna Devaraneni, Emily A. Rex, Show Ling Shyng

Research output: ResearchChapter

Abstract

Potassium channels are multimeric protein complexes regulated by diverse physiological and pharmacological ligands. The key to understanding mechanisms of channel regulation is the ability to detect structural changes associated with ligand binding. While high-resolution structural methods such as X-ray crystallography and single-particle cryo-electron microscopy offer direct visualization of channel structures, these methods do have limitations and may not be suitable for the question of interest. In this chapter, we describe the use of a photo-cross-linker unnatural amino acid, p-azido-l-phenylalanine, to probe interactions between two proteins, the sulfonylurea receptor 1 and the inwardly rectifying potassium channel Kir6.2, that form the ATP-sensitive potassium (KATP) channel complex in the absence or presence of ligands. The difference in the extent of crosslinking between a liganded state and unliganded state can be used as a readout of ligand-induced structural changes. We anticipate that the protocol described here will also be applicable for other potassium channels and protein complexes.

LanguageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages51-61
Number of pages11
Volume1684
DOIs
StatePublished - 2018

Publication series

NameMethods in Molecular Biology
Volume1684
ISSN (Print)1064-3745

Fingerprint

Phenylalanine
Adenosine Triphosphate
Ligands
Proteins
Potassium Channels
Sulfonylurea Receptors
Inwardly Rectifying Potassium Channel
Cryoelectron Microscopy
X Ray Crystallography
Potassium
Pharmacology
Amino Acids

Keywords

  • Azidophenylalanine
  • Carbamazepine
  • Glibenclamide
  • Kir6.2
  • Membrane protein
  • Pharmacological chaperone
  • Sulfonylurea receptor

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

Devaraneni, P., Rex, E. A., & Shyng, S. L. (2018). Probing Subunits Interactions in KATP Channels Using Photo-Crosslinking via Genetically Encoded p-Azido-l-phenylalanine. In Methods in Molecular Biology (Vol. 1684, pp. 51-61). (Methods in Molecular Biology; Vol. 1684). Humana Press Inc.. DOI: 10.1007/978-1-4939-7362-0_5

Probing Subunits Interactions in KATP Channels Using Photo-Crosslinking via Genetically Encoded p-Azido-l-phenylalanine. / Devaraneni, Prasanna; Rex, Emily A.; Shyng, Show Ling.

Methods in Molecular Biology. Vol. 1684 Humana Press Inc., 2018. p. 51-61 (Methods in Molecular Biology; Vol. 1684).

Research output: ResearchChapter

Devaraneni, P, Rex, EA & Shyng, SL 2018, Probing Subunits Interactions in KATP Channels Using Photo-Crosslinking via Genetically Encoded p-Azido-l-phenylalanine. in Methods in Molecular Biology. vol. 1684, Methods in Molecular Biology, vol. 1684, Humana Press Inc., pp. 51-61. DOI: 10.1007/978-1-4939-7362-0_5
Devaraneni P, Rex EA, Shyng SL. Probing Subunits Interactions in KATP Channels Using Photo-Crosslinking via Genetically Encoded p-Azido-l-phenylalanine. In Methods in Molecular Biology. Vol. 1684. Humana Press Inc.2018. p. 51-61. (Methods in Molecular Biology). Available from, DOI: 10.1007/978-1-4939-7362-0_5
Devaraneni, Prasanna ; Rex, Emily A. ; Shyng, Show Ling. / Probing Subunits Interactions in KATP Channels Using Photo-Crosslinking via Genetically Encoded p-Azido-l-phenylalanine. Methods in Molecular Biology. Vol. 1684 Humana Press Inc., 2018. pp. 51-61 (Methods in Molecular Biology).
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