Three-dimensional Reconstruction Using Transmission Electron Microscopy Reveals a Swollen, Bell-shaped Structure of Transient Receptor Potential Melastatin Type 2 Cation Channel

• Published in: The Journal of biological chemistry Vol. 282; no. 51; pp. 36961 - 36970
• Main Authors: Maruyama, Yuusuke, Ogura, Toshihiko, Mio, Kazuhiro, Kiyonaka, Shigeki, Kato, Kenta, Mori, Yasuo, Sato, Chikara
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 21.12.2007; American Society for Biochemistry and Molecular Biology
• Subjects: Adenosine Diphosphate Ribose - chemistry; Adenosine Diphosphate Ribose - metabolism; Cell Line; Cell Membrane - chemistry; Cell Membrane - metabolism; Cell Membrane - ultrastructure; Cyclic ADP-Ribose - chemistry; Cyclic ADP-Ribose - metabolism; Humans; Microscopy, Electron; Models, Molecular; Protein Structure, Tertiary - physiology; Pyrophosphatases - chemistry; Pyrophosphatases - genetics; Pyrophosphatases - metabolism; TRPC Cation Channels - agonists; TRPC Cation Channels - chemistry; TRPC Cation Channels - genetics; TRPC Cation Channels - metabolism; TRPM Cation Channels - agonists; TRPM Cation Channels - chemistry; TRPM Cation Channels - genetics; TRPM Cation Channels - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M705694200

Transient receptor potential melastatin type 2 (TRPM2) is a redox-sensitive, calcium-permeable cation channel activated by various signals, such as adenosine diphosphate ribose (ADPR) acting on the ADPR pyrophosphatase (ADPRase) domain, and cyclic ADPR. Here, we purified the FLAG-tagged tetrameric TRPM2 channel, analyzed it using negatively stained electron microscopy, and reconstructed the three-dimensional structure at 2.8-nm resolution. This multimodal sensor molecule has a bell-like shape of 18 nm in width and 25 nm in height. The overall structure is similar to another multimodal sensor channel, TRP canonical type 3 (TRPC3). In both structures, the small extracellular domain is a dense half-dome, whereas the large cytoplasmic domain has a sparse, double-layered structure with multiple internal cavities. However, a unique square prism protuberance was observed under the cytoplasmic domain of TRPM2. The FLAG epitope, fused at the C terminus of the ADPRase domain, was assigned by the antibody to a position close to the protuberance. This indicates that the agonist-binding ADPRase domain and the ion gate in the transmembrane region are separately located in the molecule.