Oligomeric state of purified transient receptor potential melastatin-1 (TRPM1), a protein essential for dim light vision

J Biol Chem. 2014 Sep 26;289(39):27019-27033. doi: 10.1074/jbc.M114.593780. Epub 2014 Aug 11.


Transient receptor potential melastatin-1 (TRPM1) is essential for the light-induced depolarization of retinal ON bipolar cells. TRPM1 likely forms a multimeric channel complex, although almost nothing is known about the structure or subunit composition of channels formed by TRPM1 or any of its close relatives. Recombinant TRPM1 was robustly expressed in insect cells, but only a small fraction was localized to the plasma membrane. Similar intracellular localization was observed when TRPM1 was heterologously expressed in mammalian cells. TRPM1 was affinity-purified from Sf9 cells and complexed with amphipol, followed by detergent removal. In blue native gels and size exclusion chromatography, TRPM1 migrated with a mobility consistent with detergent- or amphipol-bound dimers. Cross-linking experiments were also consistent with a dimeric subunit stoichiometry, and cryoelectron microscopy and single particle analysis without symmetry imposition yielded a model with approximate 2-fold symmetrical features. Finally, electron microscopy of TRPM1-antibody complexes revealed a large particle that can accommodate TRPM1 and two antibody molecules. Taken together, these data indicate that purified TRPM1 is mostly dimeric. The three-dimensional structure of TRPM1 dimers is characterized by a small putative transmembrane domain and a larger domain with a hollow cavity. Blue native gels of solubilized mouse retina indicate that TRPM1 is present in two distinct complexes: one similar in size to the recombinant protein and one much larger. Because dimers are likely not functional ion channels, these results suggest that additional partner subunits participate in forming the transduction channel required for dim light vision and the ON pathway.

Keywords: Cryoelectron Microscopy; G Protein Signaling; Ion Channel; Membrane Protein; Transient Receptor Potential Channels (TRP Channels); Vision.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Eye Proteins* / chemistry
  • Eye Proteins* / genetics
  • Eye Proteins* / isolation & purification
  • Eye Proteins* / metabolism
  • HEK293 Cells
  • Humans
  • Mice
  • Protein Multimerization / physiology*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Sf9 Cells
  • Spodoptera
  • TRPM Cation Channels* / chemistry
  • TRPM Cation Channels* / genetics
  • TRPM Cation Channels* / isolation & purification
  • TRPM Cation Channels* / metabolism
  • Vision, Ocular / physiology*


  • Eye Proteins
  • Recombinant Proteins
  • TRPM Cation Channels
  • Trpm1 protein, mouse

Associated data

  • PDB/1IGY
  • PDB/3J5P