Related Experiment Video
Updated: Mar 18, 2026

Yeast Luminometric and Xenopus Oocyte Electrophysiological Examinations of the Molecular Mechanosensitivity of TRPV4
Published on: December 31, 2013
The proposed channel-enzyme transient receptor potential melastatin 2 does not possess ADP ribose hydrolase activity
Iordan Iordanov1,2, Csaba Mihályi1,2, Balázs Tóth1,2
1Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary.
Abstract:
Transient Receptor Potential Melastatin 2 (TRPM2) is a Ca(2+)-permeable cation channel essential for immunocyte activation, insulin secretion, and postischemic cell death. TRPM2 is activated by ADP ribose (ADPR) binding to its C-terminal cytosolic NUDT9-homology (NUDT9H) domain, homologous to the soluble mitochondrial ADPR pyrophosphatase (ADPRase) NUDT9. Reported ADPR hydrolysis classified TRPM2 as a channel-enzyme, but insolubility of isolated NUDT9H hampered further investigations. Here we developed a soluble NUDT9H model using chimeric proteins built from complementary polypeptide fragments of NUDT9H and NUDT9. When expressed in E.coli, chimeras containing up to ~90% NUDT9H sequence remained soluble and were affinity-purified. In ADPRase assays the conserved Nudix-box sequence of NUDT9 proved essential for activity (kcat~4-9s(-1)), that of NUDT9H did not support catalysis. Replacing NUDT9H in full-length TRPM2 with soluble chimeras retained ADPR-dependent channel gating (K1/2~1-5 μM), confirming functionality of chimeric domains. Thus, TRPM2 is not a 'chanzyme'. Chimeras provide convenient soluble NUDT9H models for structural/biochemical studies.
More Related Videos
08:27Expression and Purification of the Human Lipid-sensitive Cation Channel TRPC3 for Structural Determination by Single-particle Cryo-electron Microscopy
Published on: January 7, 2019
10:00Controllable Ion Channel Expression through Inducible Transient Transfection
Published on: February 17, 2017
Related Concept Videos
Energy to Drive Translocation
Generally, polypeptides are unfolded by two distinct...
Mechanically-gated Ion Channels
Transducer Mechanism: Enzyme-Linked Receptors
Major types that are helpful drug targets include:
Membrane Asymmetry Regulating Transporters
Flippase
Eukaryotic flippases are type-IV P-type ATPases or P4-ATPases belonging to P-type ATPase family proteins that are membrane-bound pumps involved in the ATP-mediated transport of ions and molecules across the membrane. Flippases flip specific phospholipids from the outer to the inner leaflet of a membrane. All P4-ATPases have one...
Calmodulin-dependent Signaling
The Ca2+-CaM complex does not have enzymatic activity by itself. Instead, the complex binds downstream target proteins, including membrane proteins or enzymes,...
GPCRs Regulate Adenylyl Cylase Activity