Scientists report that they have discovered an ion channel structure that could lead to new approaches to the treatment for kidney stones. The team, which published its study (“Structural Basis of TRPV5 Channel Inhibition by Econazole Revealed by Cryo-EM”) in Nature Structural and Molecular Biology, described atomic-level details of the protein that serves as a passageway for calcium across kidney cell membranes.
“The transient receptor potential vanilloid 5 (TRPV5) channel is a member of the transient receptor potential (TRP) channel family, which is highly selective for Ca2+, that is present primarily at the apical membrane of distal tubule epithelial cells in the kidney and plays a key role in Ca2+ reabsorption. Here we present the structure of the full-length rabbit TRPV5 channel as determined using cryo-EM in complex with its inhibitor econazole. This structure reveals that econazole resides in a hydrophobic pocket analogous to that occupied by phosphatidylinositides and vanilloids in TRPV1, thus suggesting conserved mechanisms for ligand recognition and lipid binding among TRPV channels,” write the investigators.
“The econazole-bound TRPV5 structure adopts a closed conformation with a distinct lower gate that occludes Ca2+ permeation through the channel. Structural comparisons between TRPV5 and other TRPV channels, complemented with molecular dynamics (MD) simulations of the econazole-bound TRPV5 structure, allowed us to gain mechanistic insight into TRPV5 channel inhibition by small molecules.”
Approximately 80% of kidney stones are comprised of calcium salts. They are extremely painful to pass, and depending on size and location can require surgery to remove. Ion channels that span kidney cell membranes help reabsorb calcium from the urine before it can form kidney stones.
The new study is the first to show molecular details of the essential kidney calcium channel, called TRPV5, in its closed form, according to Taylor Hughes, Ph.D. candidate in the department of pharmacology at Case Western Reserve University School of Medicine, adding that the research also reveals how inhibitor molecules attach to and close the channel, leaving calcium stranded in the urine where it can form kidney stones.
“Now that we know what the protein looks like in its inhibited state, drugs can be made with the intention of modulating TRPV5 activity and potentially treating kidney stones directly,” said Hughes.
Hughes and colleagues used cryo-electron microscopy (cryo-EM) to view rabbit TRPV5 attached to its inhibitor molecule, econazole. Cryo-EM enabled the researchers to zoom in and see protein structures in atomic detail. From the new vantage point, they could identify different protein regions, including the portion that crosses kidney cell membranes, and attachment sites for molecules like econazole.
“When performing cryo-EM, we shoot electrons at our frozen protein, and this allows us to take pictures of individual protein molecules. With these pictures and advanced computer software, we are able to create 3D models of these molecules. These 3D models have the potential to be so precise that we can actually see the atoms that make up the protein,” Hughes explained.
The 3D models helped the researchers predict how TRPV5 opens and closes for the first time.
“To understand how a protein moves, we need multiple structures to compare to one another,” Hughes said. “We were able to draw conclusions about the mechanisms of action by comparing our inhibitor-bound structure to a previously published TRPV6 structure solved without an inhibitor. TRPV5 and TRPV6 are part of the same subfamily of proteins and very similar in sequence as well as structure.”
Structures for four of the six TRPV subfamily members are available at near-atomic resolution for further scientific investigation, continued Hughes. According to the researchers, future studies could include targeted therapies to modulate the protein channels in people suffering from kidney stones.
