Vascular K _ATP channel structural dynamics reveal regulatory mechanism by Mg-nucleotides

Abstract

Vascular tone is dependent on smooth muscle KATP channels comprising pore-forming Kir6.1 and regulatory SUR2B subunits, in which mutations cause Cantú syndrome. Unique among KATP isoforms, they lack spontaneous activity and require Mg-nucleotides for activation. Structural mechanisms underlying these properties are unknown. Here, we determined cryogenic electron microscopy structures of vascular KATP channels bound to inhibitory ATP and glibenclamide, which differ informatively from similarly determined pancreatic KATP channel isoform (Kir6.2/SUR1). Unlike SUR1, SUR2B subunits adopt distinct rotational “propeller” and “quatrefoil” geometries surrounding their Kir6.1 core. The glutamate/aspartate-rich linker connecting the two halves of the SUR-ABC core is observed in a quatrefoil-like conformation. Molecular dynamics simulations reveal MgADP-dependent dynamic tripartite interactions between this linker, SUR2B, and Kir6.1. The structures captured implicate a progression of intermediate states between MgADP-free inactivated, and MgADP-bound activated conformations wherein the glutamate/aspartate-rich linker participates as mobile autoinhibitory domain, suggesting a conformational pathway toward KATP channel activation.

ATP-Sensitive Potassium Channels in Migraine: Translational Findings and Therapeutic Potential

“…High concentrations of ATP will inhibit the channel, while reduced ATP levels will activate and open the channel [ 32 ]. The activity of the channel is controlled by the SUR subunits due to their NBDs, where MgATP binds to NBD2 and MgADP binds to NBD1 [ 29 , 33 ]. Phosphatidylinositol 4,5-bisphosphate (PIP 2 ) is suggested to play a role in channel activity regulation, as PIP 2 activates the channel and reduces its sensitivity to ATP, thus counteracting the inhibitory effect at ATP [ 34 , 35 , 36 ].…”

“…The K ATP channels are expressed throughout the body but the combination of the different subunits of Kir6.x and SURx vary in different tissues, such as the vascular system, neuronal system, and pancreas (see Table 1 ). The pancreatic β-cells express Kir6.2/SUR1, which control the glucose-stimulated insulin secretion (and represents the most studied channel), while the Kir6.2/SUR2A channels are the predominant form found in myocardia [ 25 , 29 , 33 ]. The vascular smooth muscle cells express Kir6.1/SUR2B and these have distinct structural features from the pancreatic Kir6.2/SUR1 isoforms, as the Kir6.1 cytoplasmic regions is placed too far from the membrane to interfere with the membrane-bound PIP 2 , which is known to activate or open the Kir6.2/SUR1 channels in pancreatic β-cells [ 33 ].…”

“…The pancreatic β-cells express Kir6.2/SUR1, which control the glucose-stimulated insulin secretion (and represents the most studied channel), while the Kir6.2/SUR2A channels are the predominant form found in myocardia [ 25 , 29 , 33 ]. The vascular smooth muscle cells express Kir6.1/SUR2B and these have distinct structural features from the pancreatic Kir6.2/SUR1 isoforms, as the Kir6.1 cytoplasmic regions is placed too far from the membrane to interfere with the membrane-bound PIP 2 , which is known to activate or open the Kir6.2/SUR1 channels in pancreatic β-cells [ 33 ]. Furthermore, Kir6.1 channels do not show spontaneous channel activity, while pancreatic and myocardial Kir6.2 channels open spontaneously when ATP levels are low or absent [ 33 , 38 , 43 ].…”

Structural insights into the mechanism of nucleotide regulation of pancreatic KATP channel

“…KNtp not only enhances the ATP sensitivity of Kir6.2 but also mediates the inhibition of insulin secretagogue in the absence of nucleotides, possibly by binding to the central vestibule of SUR and restraining the mobility of Kir6 CTD (Fig. 6a) 29,30 . In the H175K cryo-EM construct, the KNtp is covalently fused to the C-terminus of SUR1 and therefore could not bind inside the SUR1 central vestibule anymore 26,27 .…”

“…The SUR subunits bind activating Mg-ADP and drugs, including insulin secretagogues and K ATP openers 11,21 . Recent advances in cryo-EM structure determination of K ATP channels in the presence of different ligand combinations by three groups have provided instrumental information about how K ATP channels are assembled from individual subunits, how inhibitory ATP binds the channel, and how chemically distinct insulin secretagogues bind at SUR subunits [22][23][24][25][26][27][28][29][30] . Moreover, the conformational changes of the SUR1 subunit upon Mg-nucleotide binding have been visualized 22,26 .…”