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Photoaffinity Labeling of RARs and Mapping of Labeled Sites by an Endoproteinase Combination Technique

  • Yuichi Hashimoto
  • Toru Sasaki
Part of the Methods in Molecular Biology book series (MIMB, volume 89)

Abstract

Photoaffinity labeling and mapping of the labeled site(s) are a superior and convenient methodology for initial structural investigation of ligand-receptor complexes. The method we describe in this chapter consists of five steps.
  1. 1.

    Design and synthesis of a photoreactive probe for retinoic acid receptor (RAR)-labeling

     
  2. 2.

    Photoaffmlty labeling of RAR

     
  3. 3.

    Sequential digestion of the labeled RAR with endoproteinases

     
  4. 4

    High-performance liquid chromatography (HPLC) analysis of the digests to determine the amino-acid sequence context of the labeled sites (endoproteinase combination technique).

     
  5. 5.

    Mapping of the labeled site(s) by comparison of the determined context with the known amino-acid sequence of the RAR</lt>

     

Keywords

Retinoic Acid Receptor Thionyl Chloride Fluorescent Peak Incubation Buffer Dialysis Buffer 
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.

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Copyright information

© Humana Press Inc, Totowa, NJ 1998

Authors and Affiliations

  • Yuichi Hashimoto
    • 1
  • Toru Sasaki
    • 1
  1. 1.Institute of Molecular and Cellular BiosciencesUniversity of TokyoJapan

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