1-20 of 31270
Save search
Follow your search
Access your saved searches in your account
Would you like to receive an alert when new items match your search?
Close Modal
Sort by
Journal Articles

On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC

Hridya Valia Madapally, Adel Hussein, Martin Wazar Eriksen, Bjørn Panyella Pedersen, David L. Stokes, Himanshu Khandelia
Journal: Journal of General Physiology
J Gen Physiol (2025) 158 (1): e202513794.
https://doi.org/10.1085/jgp.202513794
Published: 12 December 2025
View article titled, On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC
Open the PDF for On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC in another window
Includes: Supplementary data
Images
Transport mechanism through KdpFABC.(A) Schematic illustration of KdpFABC in three catalytic states. In the E2 state (left), a K+ ion from the periplasm is bound to the selectivity filter in KdpA and the tunnel is blocked at the interface between KdpA and KdpB. In the ATP bound E1 state (middle), the tunnel is open and K+ has access to the canonical binding site (CBS) in KdpB. In the E2-P state (right), in which Asp307 is transiently phosphorylated, the tunnel is again blocked, but the ion has been moved to the release site in KdpB. After hydrolyzing the aspartyl phosphate and releasing the K+ to the cytoplasm, the cycle resets. The high-energy E1∼P state is not shown. KdpA is colored green. The transmembrane domain of KdpB is brown, and the cytoplasmic domains are yellow (A-domain), red (N-domain), and blue (P-domain). KdpC is purple and KdpF is not shown. (B) Closeup of the tunnel showing alternative scenarios in which it is filled either with water (as seen for example in PDB 7LC3) or with K+ ions (as seen for example in PDB 7ZRK). In the former scenario, an individual K+ moves through the tunnel to reach the CBS, whereas the latter scenario would presumably involve a relay. S3 denotes the primary binding site for K+ in the selectivity filter. Phe232 from KdpB resides at the subunit interface and has been shown to play a role in coupling ATPase activity to transport (Silberberg et al., 2021). Lys586 resides in the CBS and has been hypothesized to play a role in moving K+ into the release site (Sweet et al., 2021). M4 refers to a transmembrane helix in KdpB that is unwound in the vicinity of the conserved Pro264, thus forming the primary binding site in KdpB.

Transport mechanism through KdpFABC. (A) Schematic illustration of KdpFAB...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
Figure 1. Transport mechanism through KdpFABC. (A) Schematic illustration of KdpFABC in three catalytic states. In the E2 state (left), a K+ ion from the periplasm is bound to the selectivity filter in KdpA and the tunnel is blocked at the More about this image found in Transport mechanism through KdpFABC. (A) Schematic illustration of KdpFAB...
Images
K+ locations with 1 or 2 K+ ions in the tunnel.(A and C) Distance between the K+ ions and the center of mass of the CBS in systems with 1 and 2 K+ ions, respectively, placed initially in the translocation passage. (B and D) The red spheres represent the oxygen atoms that form the CBS (in B one of the oxygen atoms is behind the white sphere) and the white sphere represents the center of mass of those oxygen atoms. The purple spheres represent K+ ions at S3 and I1. KdpA, KdpB, KdpC, and KdpF are shown in green, brown, purple, and cyan, respectively. Selectivity filter loops are depicted in dark green color.

K+ locations with 1 or 2 K+ ions in the tunnel. (A a...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
Figure 2. K+ locations with 1 or 2 K+ ions in the tunnel. (A and C) Distance between the K+ ions and the center of mass of the CBS in systems with 1 and 2 K+ ions, respectively, placed initially in the translocation passage. (B and D) The red More about this image found in K+ locations with 1 or 2 K+ ions in the tunnel. (A a...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
More about this image found in
Images
Number of K+ions that remain in the translocation passage in simulations starting with different initial numbers of K+ions.

Number of K + ions that remain in the translocation passage in...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
Figure 3. Number of K + ions that remain in the translocation passage in simulations starting with different initial numbers of K + ions. More about this image found in Number of K + ions that remain in the translocation passage in...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
More about this image found in
Images
Energetics of K+ transport.(A) Free energy of passage of K+ along the tunnel obtained using multiple-walker well-tempered Metadynamics. d.x ˜ −0.20 nm corresponds to the ion-binding site, CBS in KdpB, d.x ≈ 3.45 nm corresponds to the S3 site of KdpA and d.x ≈ 2.65 nm corresponds to the I1 site at KdpA. The I1 site has been chosen as the zero of the free energy. The light blue shaded region corresponds to the transition state region. (B) The transition state at d.x ≈ 0.60 nm. The K+ ion is at the interface and is yet to cross Phe232. (C) K+ ion at the B site coordinated by Asn624, Asp583, Cys261, and Ile263.

Energetics of K+ transport . (A) Free energy of passage of K...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
Figure 4. Energetics of K+ transport . (A) Free energy of passage of K+ along the tunnel obtained using multiple-walker well-tempered Metadynamics. d.x ˜ −0.20 nm corresponds to the ion-binding site, CBS in KdpB, d.x ≈ 3.45 nm corresponds to More about this image found in Energetics of K+ transport . (A) Free energy of passage of K...
Images
Hydration of the tunnel.(A) Number of water molecules present in the tunnel when one K+ or three K+ ions are initially placed in the tunnel. (B) The probability density function (PDF) of water molecules along the axis of the tunnel. The tunnel region was identified using HOLE. The data were normalized so that the integral represents the average number of water molecules.

Hydration of the tunnel. (A) Number of water molecules present in the tun...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
Figure 5. Hydration of the tunnel. (A) Number of water molecules present in the tunnel when one K+ or three K+ ions are initially placed in the tunnel. (B) The probability density function (PDF) of water molecules along the axis of the More about this image found in Hydration of the tunnel. (A) Number of water molecules present in the tun...
Images
F232A mutant uncouples ATPase from potassium transport. (A) ATPase activity for WT as well as KdpB-F232I and KdpB-F232A mutations. Each of these KdpB mutants was tested with a WT selectivity filter (KdpA-WT) and a Q116R selectivity filter (KdpA-Q116R) in the presence (+) and absence (−) of 150 mM K+. (B) Transport of WT and the two KdpB-F232 mutants measured with SSME. Uncoupling by the F232A mutation is evident from the K+-independent ATPase activity and the complete lack of transport. The F232I mutant retains K+ dependence and coupling, but exhibits lower ATPase and transport activities. Raw data from the SSME transport assay are shown in Fig. S13.

F232A mutant uncouples ATPase from potassium transport. (A) ATPase activit...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
Figure 6. F232A mutant uncouples ATPase from potassium transport. (A) ATPase activity for WT as well as KdpB-F232I and KdpB-F232A mutations. Each of these KdpB mutants was tested with a WT selectivity filter (KdpA-WT) and a Q116R selectivity More about this image found in F232A mutant uncouples ATPase from potassium transport. (A) ATPase activit...
Images
Anomalous signal derived from X-ray crystallography of KdpFABC. Panel A shows an overview of the anomalous signal in KdpFABC. This represents one of three copies in the asymmetric unit (chains A–D), with the others shown in Fig. S14. The model corresponds to the previous X-ray structure (PDB accession no. 5MRW) which was used to phase the current data by molecular replacement. The only significant signal was observed in the S3 position of the selectivity filter, shown in closeup in panel B. The mesh surface corresponds to 4σ with the peak slightly exceeding 7σ. No significant signal was seen within the tunnel connecting the subunits. KdpA is colored green, KdpB brown, and KdpC is purple.

Anomalous signal derived from X-ray crystallography of KdpFABC. Panel A sh...

in On the mechanism of K+ transport through the inter-subunit tunnel of KdpFABC > Journal of General Physiology
Published: 12 December 2025
Figure 7. Anomalous signal derived from X-ray crystallography of KdpFABC. Panel A shows an overview of the anomalous signal in KdpFABC. This represents one of three copies in the asymmetric unit (chains A–D), with the others shown in Fig. S14 . More about this image found in Anomalous signal derived from X-ray crystallography of KdpFABC. Panel A sh...
Journal Articles

Towards a unified gating scheme for the CNBD ion channel family

Jenna L. Lin, Baron Chanda
Journal: Journal of General Physiology
J Gen Physiol (2025) 158 (1): e202513849.
https://doi.org/10.1085/jgp.202513849
Published: 11 December 2025
View article titled, Towards a unified gating scheme for the CNBD ion channel family
Open the PDF for Towards a unified gating scheme for the CNBD ion channel family in another window
Includes: Supplementary data
Images
Phenotype chart of HCN-EAG chimeras illustrates the design principles for hyperpolarization-dependent gating. (A) Schematic showing the three structural modules that contribute to hyperpolarization-dependent gating (left). Simplified representations of HCN1 (center) and EAG (right) highlighting the corresponding modules. (B–G) Gating phenotypes of various chimeras and mutants. (B) HHHHE, (C) HHHEH, (D) HHHEE, (E) EEEHH, described in Cowgill et al. (2019); (F) HHHEΔC, described in Lin et al. (2024); and (G) HCN1 C-terminal deletion, from (Wainger et al, 2001) and (Wang et al, 2001). This phenotype chart shows that the HCN-derived VSD is necessary but not sufficient for hyperpolarization-dependent gating. In addition, at least one of the two secondary structural modules—either the PD or the CTD—must also be derived from HCN to confer this gating behavior. When all three modules are appropriately matched, as in wild-type HCN channels, the system exhibits more robust hyperpolarization-dependent gating. VSD, voltage-sensing domain; PD, pore domain; CTD, C-terminal domain.

Phenotype chart of HCN-EAG chimeras illustrates the design principles for h...

in Towards a unified gating scheme for the CNBD ion channel family > Journal of General Physiology
Published: 11 December 2025
Figure 1. Phenotype chart of HCN-EAG chimeras illustrates the design principles for hyperpolarization-dependent gating. (A) Schematic showing the three structural modules that contribute to hyperpolarization-dependent gating (left). Simplified More about this image found in Phenotype chart of HCN-EAG chimeras illustrates the design principles for h...
Images
Allosteric factor, n, of the inverted coupling model modulates gating polarity. (A) Gating scheme of the inverted coupling model. C and O are when the pore is closed and opened, respectively. VD and VH are the voltage sensor upon membrane depolarization and upon membrane hyperpolarization, respectively. K1 is a voltage-dependent equilibrium state constant for movement of the voltage sensor. K2 is voltage-independent for opening and closure of the pore. n is the allosteric factor. Equations for calculating the PO−V curves are described in Materials and methods. (B) Family of PO−V plots based on the gating scheme in A. The K10 = 1, q1 = −1, and K2 = 1 are the same for all except for n, which is varied as shown in the legend. (C)PO−V scatter plot of hEAG (adapted from Cowgill et al. [2019]). These data were fitted (dashed black line) with the following parameters: K10 = 22.4, q1 = −1.46, and K2 = 143. n = 3.38 × 10−5. PO-V plots in blue correspond to varying values of n when n is <1. (D)PO−V scatter plot of mHCN1 (adapted from Cowgill et al. [2019]). These data were fitted (dashed black line) with the following parameters: K10 = 1.13 × 10−4, q1 = −2.03, and K2 = 0.0139, n = 1,550. PO−V plots in red correspond to varying values of n when n is >1.

Allosteric factor, n, of the inverted coupling model modul...

in Towards a unified gating scheme for the CNBD ion channel family > Journal of General Physiology
Published: 11 December 2025
Figure 2. Allosteric factor, n, of the inverted coupling model modulates gating polarity. (A) Gating scheme of the inverted coupling model. C and O are when the pore is closed and opened, respectively. VD and VH are the voltage sensor upon More about this image found in Allosteric factor, n, of the inverted coupling model modul...
Images
Bipolar gating phenotype is described using the three-state gating polarity model. (A) Gating scheme of the three-state gating polarity model. C is when the channel is closed. OH is channel opening upon membrane hyperpolarization, and OD is channel opening upon depolarization. K1 and K2 are voltage-dependent equilibrium state constants. Equations for calculating PO−V curves are described in Materials and methods. All plots were generated using the parameters q1 = −2 and q2 = 2. (B)PO−V scatter plot of D540K-hERG mutant (adapted from Tristani-Firouzi et al. [2002]). These data were normalized again such that the maximum PO is normalized to 1 (i.e., relative current/maximum relative current). The bipolar gating phenotype is observed when K10 = K20 = 0.025 (dashed black line). Fitting of these data can be approximated by setting K10 < K20 such that K10 = 0.0001 and K20 = 5 (solid black line). (C) Series of PO−V plots show the result of K10 becoming increasingly smaller than K20 (solid lines with increasingly lighter shades of red), where K20 = 0.025 and K10 is equal to K20 multiplied by a varying factor (shown in figure legend). (D) Series of PO−V plots show the result of K20 becoming increasingly smaller than K10 (solid lines with increasingly lighter shades of blue), where K10 = 0.025 and K20 is equal to K10 multiplied by a varying factor (shown in figure legend).

Bipolar gating phenotype is described using the three-state gating polarity...

in Towards a unified gating scheme for the CNBD ion channel family > Journal of General Physiology
Published: 11 December 2025
Figure 3. Bipolar gating phenotype is described using the three-state gating polarity model. (A) Gating scheme of the three-state gating polarity model. C is when the channel is closed. OH is channel opening upon membrane hyperpolarization, and More about this image found in Bipolar gating phenotype is described using the three-state gating polarity...
Images
Unconstrained five-state gating polarity model. (A) Gating scheme of the five-state gating polarity model. Voltage-dependent transition steps, K1 and K2, and voltage-independent transition steps, K3 and K4, are all freely floating parameters. C is the closed state, and O is the open state of the pore. V is the state of the voltage sensor, where VR is at rest. Subscripts H and D for all O and V states indicate states upon membrane hyperpolarization and upon membrane depolarization, respectively. Equations for calculating PO−V plots are described in Materials and methods. Fitting of the PO−V plots to data using the gating scheme are shown as a solid black line in figure panels (B–I). Parameter values used for these fittings are reported in Table S1. PO−V scatter plot data in B and C are adapted from Tristani-Firouzi et al. (2002), and normalized again such that the maximum PO is normalized to 1 (i.e., relative current/maximum relative current). PO−V scatter plot data in D–I are adapted from Lin et al. (2024). (B–I)PO−V scatter plot data of D540K-Q664A hERG (upward-pointing triangle ▲), D540K-L666A hERG (square ■), HHHEH (diamond ♦), HHHEH2 (downward-pointing triangle ▼), HHHES (asterisk *), HHHER (right-pointing triangle ►), HHHEK (pentagram ★), and HHHEA (left-pointing triangle ◄).

Unconstrained five-state gating polarity model. (A) Gating scheme of the f...

in Towards a unified gating scheme for the CNBD ion channel family > Journal of General Physiology
Published: 11 December 2025
Figure 4. Unconstrained five-state gating polarity model. (A) Gating scheme of the five-state gating polarity model. Voltage-dependent transition steps, K1 and K2, and voltage-independent transition steps, K3 and K4, are all freely floating More about this image found in Unconstrained five-state gating polarity model. (A) Gating scheme of the f...
Images
Five-state gating polarity model does not fit conditions where VSD is constant. (A) Gating scheme of the five-state gating polarity model. Voltage-dependent steps, K1 and K2, are bolded to indicate constrained values (i.e., K1 and K2 parameter values used for fitting HHHEH are applied to all HHHE-X chimeras). K3 and K4 are voltage-independent transition steps and vary in each chimera. Abbreviations for each state and equations for calculations of open probabilities are the same as Fig. 4 A. Fitting of PO−V scatter plot data to the gating scheme described in A is shown in the following panels as a solid black line. Parameter values used in these fittings are in Table S2. PO−V scatter plot data are adapted from Lin et al. (2024). (B–G)PO−V scatter plot of HHHEH (diamond ♦), HHHEH2 (downward-pointing triangle ▼), HHHES (asterisk *), HHHER (right-pointing triangle ►), HHHEK (pentagram ★), and HHHEA (left-pointing triangle ◄). (i–iii) Collection of PO−V plots in F and G, such that K3 = 1 × 10−7 and K4 is varied to the following parameter values: (i) K4 = 1, (ii) K4 = 10, and (iii) K4 = 100.

Five-state gating polarity model does not fit conditions where VSD is const...

in Towards a unified gating scheme for the CNBD ion channel family > Journal of General Physiology
Published: 11 December 2025
Figure 5. Five-state gating polarity model does not fit conditions where VSD is constant. (A) Gating scheme of the five-state gating polarity model. Voltage-dependent steps, K1 and K2, are bolded to indicate constrained values (i.e., K1 and K2 More about this image found in Five-state gating polarity model does not fit conditions where VSD is const...
Images
Seven-state gating polarity model describes bipolar gating phenotype of HHHE-X chimeras. (A) Gating scheme of the seven-state gating polarity model. Voltage-dependent steps, K1 and K2, are bolded to indicate they are constrained, where K1 and K2 values from HHHEH fitting are applied to all HHHE-X chimeras. K3, K4, K5, and K6 are voltage-independent transition steps and vary for each chimera. C, OH, and OD are the closed and opened states of the pore, where OH is for hyperpolarization and OD is for depolarization. V is the state of the voltage sensor such that VR is at rest, VH is upon membrane hyperpolarization, and VD is upon membrane depolarization. N is the interaction due to the cytosolic C terminus upon hyperpolarization, NH, and depolarization, ND. Equations for open probability calculations are in Materials and methods, and parameter values used for fitting are reported in Table S3. (B–J)PO−V plots of fitting the gating scheme in A to the PO−V scatter plot data are shown as a solid black line in each figure panel. PO−V scatter plot data from hERG mutants, i.e., B–D, are adapted from Tristani-Firouzi et al. (2002). These data have been normalized again such that the maximum PO value across all test potentials is normalized to 1 (i.e., relative current/maximum relative current). PO−V scatter plot data from HHHE-X, i.e., E–J, are adapted from Lin et al. (2024). PO−V scatter plot data of D540K hERG (circle ●), D540K-Q664A hERG (upward-pointing triangle ▲), D540K-L666A hERG (square ■), HHHEH (diamond ♦), HHHEH2 (downward-pointing triangle ▼), HHHES (asterisk *), HHHER (right-pointing triangle ►), HHHEK (pentagram ★), and HHHEA (left-pointing triangle ◄).

Seven-state gating polarity model describes bipolar gating phenotype of HHH...

in Towards a unified gating scheme for the CNBD ion channel family > Journal of General Physiology
Published: 11 December 2025
Figure 6. Seven-state gating polarity model describes bipolar gating phenotype of HHHE-X chimeras. (A) Gating scheme of the seven-state gating polarity model. Voltage-dependent steps, K1 and K2, are bolded to indicate they are constrained, where More about this image found in Seven-state gating polarity model describes bipolar gating phenotype of HHH...
Images
Physical interpretation of the seven-state gating polarity model for CNBD channels. This cartoon depicts our hypothesis of the underlying mechanism in the gating scheme of Fig. 6 A. The shaded circle represents the pore in the closed state, while the blank circle is in the opened state. Distinct orientations of the CNBD are drawn to represent different interactions between the cytosolic C-terminal domain and the transmembrane domain regions.

Physical interpretation of the seven-state gating polarity model for CNBD c...

in Towards a unified gating scheme for the CNBD ion channel family > Journal of General Physiology
Published: 11 December 2025
Figure 7. Physical interpretation of the seven-state gating polarity model for CNBD channels. This cartoon depicts our hypothesis of the underlying mechanism in the gating scheme of Fig. 6 A . The shaded circle represents the pore in the closed More about this image found in Physical interpretation of the seven-state gating polarity model for CNBD c...
Journal Articles

Novel binding mode for negative allosteric NMDA receptor modulators

James S. Lotti, Jed T. Syrenne, Avery J. Benton, Ahmad Al-Mousawi, Lauren E. Cornelison, Christopher J. Trolinder, Feng Yi, Zhucheng Zhang, Cindee K. Yates-Hansen, Levi J. McClelland, James Bosco, Andrew R. Rau, Rasmus P. Clausen, Kasper B. Hansen
Journal: Journal of General Physiology
J Gen Physiol (2025) 158 (1): e202513872.
https://doi.org/10.1085/jgp.202513872
Published: 03 December 2025
View article titled, Novel binding mode for negative allosteric NMDA receptor modulators
Open the PDF for Novel binding mode for negative allosteric NMDA receptor modulators in another window
Includes: Supplementary data
Images
Evaluation of NAMs at NMDA receptor subtypes. (A) Chemical structures of NAMs, UCM-101, TCN-213, TCN-201, MPX-004, and MPX-007. (B) Representative two-electrode voltage-clamp recordings from recombinant GluN1/2A or GluN1/2B receptors activated by the indicated concentration of glycine in the continuous presence of 100 µM glutamate and inhibited by increasing concentrations of TCN-213 or UCM-101. (C) Concentration–response data for TCN-213, UCM-101, and TCN-201 at NMDA receptor subtypes activated by 1 µM glycine in the continuous presence of 100 µM glutamate. (D) Concentration–response data for TCN-213, UCM-101, and TCN-201 at NMDA receptor subtypes activated by a glycine concentration close to the glycine EC50 in the continuous presence of 100 µM glutamate (1 µM glycine for GluN1/2A, 0.3 µM for GluN1/2B and GluN1/2C, and 0.1 µM for GluN1/2D). A dashed line indicates data for GluN1/2A from (C). See Tables 1 and 2 for NAM IC50 and glycine EC50 values.

Evaluation of NAMs at NMDA receptor subtypes. (A) Chemical structures of N...

in Novel binding mode for negative allosteric NMDA receptor modulators > Journal of General Physiology
Published: 03 December 2025
Figure 1. Evaluation of NAMs at NMDA receptor subtypes. (A) Chemical structures of NAMs, UCM-101, TCN-213, TCN-201, MPX-004, and MPX-007. (B) Representative two-electrode voltage-clamp recordings from recombinant GluN1/2A or GluN1/2B receptors More about this image found in Evaluation of NAMs at NMDA receptor subtypes. (A) Chemical structures of N...